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CONTENTS i
FOURTEENTH EDITION
Junqueira’s
Basic Histology T E X T A N D AT L A S
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Contents
KEY FEATURES╇ VI╇ |╇ PREFACE╇IX╇|╇ACKNOWLEDGMENTS╇XI
iii
iv CONTENTS
ix
Acknowledgments
I wish to thank the students at Indiana University School of these valuable resources is gratefully acknowledged. Students are
Medicine and the undergraduates at Indiana University with referred to those review books for hundreds of additional self-
whom I have studied histology and cell biology for over 30 years assessment questions.
and from whom I have learned much about presenting basic I am also grateful to my colleagues and reviewers from
concepts most effectively. Their input has greatly helped in the throughout the world who provided specialized expertise
task of maintaining and updating the presentations in this classic or original photographs, which are also acknowledged in
textbook. As with the last edition the help of Sue Childress and figure captions. I thank those professors and students in the
Dr. Mark Braun was invaluable in slide preparation and the United States, as well as Argentina, Canada, Iran, Ireland, Italy,
virtual microscope for human histology respectively. Pakistan, and Syria, who provided useful suggestions that
A major change in this edition is the inclusion of self- have improved the new edition of Junqueira’s Basic Histology.
assessment questions with each topic/chapter. Many of these Finally, I am pleased to acknowledge the help and collegiality
questions were used in my courses, but others are taken or provided by the staff of McGraw-Hill, especially editors
modified from a few of the many excellent review books published Michael Weitz and Brian Kearns, whose work made possible
by McGraw-Hill/Lange for students preparing to take the U.S. publication of this 14th edition of Junqueira’s Basic Histology.
Medical Licensing Examination. These include Histology and Cell
Biology: Examination and Board Review, by Douglas Paulsen; Anthony L. Mescher
USMLE Road Map: Histology, by Harold Sheedlo; and Anatomy,
Indiana University School of Medicine
Histology, & Cell Biology: PreTest Self-Assessment & Review, by
Robert Klein and George Enders. The use here of questions from mescher@indiana.edu
xi
C H A P T E R
1
PREPARATION OF TISSUES FOR STUDY
Histology & Its
Methods of Study
1 AUTORADIOGRAPHY 9
Fixation 1 CELL & TISSUE CULTURE 10
Embedding & Sectioning 3
ENZYME HISTOCHEMISTRY 10
Staining 3
LIGHT MICROSCOPY 4 VISUALIZING SPECIFIC MOLECULES 10
Bright-Field Microscopy 4 Immunohistochemistry 11
Fluorescence Microscopy 5 Hybridization Techniques 12
Phase-Contrast Microscopy 5 INTERPRETATION OF STRUCTURES IN TISSUE
Confocal Microscopy 5 SECTIONS 14
Polarizing Microscopy 7 SUMMARY OF KEY POINTS 15
ELECTRON MICROSCOPY 8 ASSESS YOUR KNOWLEDGE 16
Transmission Electron Microscopy 8
Scanning Electron Microscopy 9
1
2 CHAPTER 1â•… ■â•… Histology & Its Methods of Study
52°- 60°C
Drive wheel
Block holder
Paraffin block
Tissue
Steel knife
Most tissues studied histologically are prepared as shown, with Similar steps are used in preparing tissue for transmission elec-
this sequence of steps (a): tron microscopy (TEM), except special fixatives and dehydrating
solutions are used with smaller tissue samples and embedding
■⌀ Fixation: Small pieces of tissue are placed in solutions of
involves epoxy resins which become harder than paraffin to allow
chemicals that cross-link proteins and inactivate degradative
very thin sectioning.
enzymes, which preserves cell and tissue structure.
■⌀ Dehydration: The tissue is transferred through a series of (b) A microtome is used for sectioning paraffin-embedded tissues
increasingly concentrated alcohol solutions, ending in 100%, for light microscopy. The trimmed tissue specimen is mounted
which removes all water. in the paraffin block holder, and each turn of the drive wheel by
■⌀ Clearing: Alcohol is removed in organic solvents in which the histologist advances the holder a controlled distance, gener-
both alcohol and paraffin are miscible. ally from 1 to 10 μm. After each forward move, the tissue block
■⌀ Infiltration: The tissue is then placed in melted paraffin until it passes over the steel knife edge and a section is cut at a thickness
becomes completely infiltrated with this substance. equal to the distance the block advanced. The paraffin sections
■⌀ Embedding: The paraffin-infiltrated tissue is placed in a small are placed on glass slides and allowed to adhere, deparaffinized,
mold with melted paraffin and allowed to harden. and stained for light microscope study. For TEM, sections less than
■⌀ Trimming: The resulting paraffin block is trimmed to expose 1 μm thick are prepared from resin-embedded cells using an
the tissue for sectioning (slicing) on a microtome. ultramicrotome with a glass or diamond knife.
organs are placed as soon as possible after removal from the microscopy, react with the amine groups (NH2) of proteins,
body in solutions of stabilizing or cross-linking compounds preventing their degradation by common proteases. Glutaral-
called fixatives. Because a fixative must fully diffuse through dehyde also cross-links adjacent proteins, reinforcing cell and
the tissues to preserve all cells, tissues are usually cut into small ECM structures.
fragments before fixation to facilitate penetration. To improve Electron microscopy provides much greater magnification
cell preservation in large organs fixatives are often introduced and resolution of very small cellular structures and fixation
via blood vessels, with vascular perfusion allowing fixation must be done very carefully to preserve additional “ultra-
rapidly throughout the tissues. structural” detail. Typically in such studies glutaraldehyde-
One widely used fixative for light microscopy is forma- treated tissue is then immersed in buffered osmium
lin, a buffered isotonic solution of 37% formaldehyde. Both tetroxide, which preserves (and stains) cellular lipids as well
this compound and glutaraldehyde, a fixative used for electron as proteins.
Preparation of Tissues for Study 3
C H A P T E R
To permit thin sectioning fixed tissues are infiltrated and Most cells and extracellular material are completely color-
embedded in a material that imparts a firm consistency. less, and to be studied microscopically tissue sections must
Embedding materials include paraffin, used routinely for light be stained (dyed). Methods of staining have been devised that
microscopy, and plastic resins, which are adapted for both make various tissue components not only conspicuous but also
light and electron microscopy. distinguishable from one another. Dyes stain material more or
Before infiltration with such media the fixed tissue must less selectively, often behaving like acidic or basic compounds
undergo dehydration by having its water extracted gradually and forming electrostatic (salt) linkages with ionizable radicals
1
by transfers through a series of increasing ethanol solutions, of macromolecules in tissues. Cell components such as nucleic
Histology & Its Methods of Study ╇ ■╇ Preparation of Tissues for Study
ending in 100% ethanol. The ethanol is then replaced by an acids with a net negative charge (anionic) have an affinity for
organic solvent miscible with both alcohol and the embedding basic dyes and are termed basophilic; cationic components,
medium, a step referred to as clearing because infiltration with such as proteins with many ionized amino groups, stain more
the reagents used here gives the tissue a translucent appearance. readily with acidic dyes and are termed acidophilic.
The fully cleared tissue is then placed in melted paraffin Examples of basic dyes include toluidine blue, alcian blue,
in an oven at 52°-60°C, which evaporates the clearing solvent and methylene blue. Hematoxylin behaves like a basic dye,
and promotes infiltration of the tissue with paraffin, and then staining basophilic tissue components. The main tissue com-
embedded by allowing it to harden in a small container of ponents that ionize and react with basic dyes do so because of
paraffin at room temperature. Tissues to be embedded with acids in their composition (DNA, RNA, and glycosaminogly-
plastic resin are also dehydrated in ethanol and then infiltrated cans). Acid dyes (eg, eosin, orange G, and acid fuchsin) stain
with plastic solvents that harden when cross-linking polymer- the acidophilic components of tissues such as mitochondria,
izers are added. Plastic embedding avoids the higher tempera- secretory granules, and collagen.
tures needed with paraffin, which helps avoid tissue distortion. Of all staining methods, the simple combination of
The hardened block with tissue and surrounding embed- hematoxylin and eosin (H&E) is used most commonly.
ding medium is trimmed and placed for sectioning in an Hematoxylin stains DNA in the cell nucleus, RNA-rich por-
instrument called a microtome (Figure 1–1). Paraffin sections tions of the cytoplasm, and the matrix of cartilage, produc-
are typically cut at 3-10 μm thickness for light microscopy, but ing a dark blue or purple color. In contrast, eosin stains other
electron microscopy requires sections less than 1 μm thick. cytoplasmic structures and collagen pink (Figure 1–2a). Here
One micrometer (1 μm) equals 1/1000 of a millimeter (mm) eosin is considered a counterstain, which is usually a single
or 10–6 m. Other spatial units commonly used in microscopy dye applied separately to distinguish additional features of a
are the nanometer (1 nm = 0.001 μm = 10–6 mm = 10–9 m) and tissue. More complex procedures, such as trichrome stains (eg,
angstrom (1 Å = 0.1 nm or 10–4 μm). The sections are placed Masson trichrome), allow greater distinctions among various
on glass slides and stained for light microscopy or on metal extracellular tissue components.
grids for electron microscopic staining and examination. The periodic acid-Schiff (PAS) reaction utilizes the
hexose rings of polysaccharides and other carbohydrate-rich
› ╺╺ MEDICAL APPLICATION tissue structures and stains such macromolecules distinctly
purple or magenta. Figure 1–2b shows an example of cells with
Biopsies are tissue samples removed during surgery or rou- carbohydrate-rich areas well-stained by the PAS reaction. The
tine medical procedures. In the operating room, biopsies DNA of cell nuclei can be specifically stained using a modifica-
are fixed in vials of formalin for processing and microscopic tion of the PAS procedure called the Feulgen reaction.
analysis in a pathology laboratory. If results of such analyses Basophilic or PAS-positive material can be further identi-
are required before the medical procedure is completed, for fied by enzyme digestion, pretreatment of a tissue section with
example to know whether a growth is malignant before the an enzyme that specifically digests one substrate. For example,
patient is closed, a much more rapid processing method is pretreatment with ribonuclease will greatly reduce cytoplas-
used. The biopsy is rapidly frozen in liquid nitrogen, preserv- mic basophilia with little overall effect on the nucleus, indicat-
ing cell structures and making the tissue hard and ready for ing the importance of RNA for the cytoplasmic staining.
sectioning. A microtome called a cryostat in a cabinet at Lipid-rich structures of cells are revealed by avoiding the
subfreezing temperature is used to section the block with processing steps that remove lipids, such as treatment with
tissue, and the frozen sections are placed on slides for rapid heat and organic solvents, and staining with lipid-soluble
staining and microscopic examination by a pathologist. dyes such as Sudan black, which can be useful in diagnosis
Freezing of tissues is also effective in histochemical stud- of metabolic diseases that involve intracellular accumulations
ies of very sensitive enzymes or small molecules because of cholesterol, phospholipids, or glycolipids. Less common
freezing, unlike fixation, does not inactivate most enzymes. methods of staining can employ metal impregnation tech-
Finally, because clearing solvents often dissolve cell lipids in niques, typically using solutions of silver salts to visual certain
fixed tissues, frozen sections are also useful when structures ECM fibers and specific cellular elements in nervous tissue.
containing lipids are to be studied histologically. The Appendix lists important staining procedures used for
most of the light micrographs in this book.
4 CHAPTER 1â•… ■â•… Histology & Its Methods of Study
FIGURE 1–2╇ Hematoxylin and eosin (H&E) and periodic acid-Schiff (PAS) staining.
G G
G
L
L
a b
Micrographs of epithelium lining the small intestine, (a) stained lumen, where projecting microvilli have a prominent layer of
with H&E, and (b) stained with the PAS reaction for glycoproteins. glycoproteins at the lumen (L) and in the mucin-rich secretory
With H&E, basophilic cell nuclei are stained purple while cyto- granules of goblet cells. Cell surface glycoproteins and mucin are
plasm stains pink. Cell regions with abundant oligosaccharides PAS-positive because of their high content of oligosaccharides
on glycoproteins, such as the ends of the cells at the lumen (L) and polysaccharides respectively. The PAS-stained tissue was
or the scattered mucus-secreting goblet cells (G), are poorly counterstained with hematoxylin to show the cell nuclei. (a. X400;
stained. With PAS, however, cell staining is most intense at the b. X300)
Slide preparation, from tissue fixation to observation (or ocular lens) further magnifying this image and projecting
with a light microscope, may take from 12 hours to 2½ days, it onto the viewer’s retina or a charge-coupled device (CCD)
depending on the size of the tissue, the embedding medium, highly sensitive to low light levels with a camera and monitor.
and the method of staining. The final step before microscopic The total magnification is obtained by multiplying the magni-
observation is mounting a protective glass coverslip on the fying power of the objective and ocular lenses.
slide with clear adhesive. The critical factor in obtaining a crisp, detailed image
with a light microscope is its resolving power, defined as the
smallest distance between two structures at which they can be
›â•ºLIGHT MICROSCOPY seen as separate objects. The maximal resolving power of the
light microscope is approximately 0.2 μm, which can permit
Conventional bright-field microscopy, as well as more special- clear images magnified 1000-1500 times. Objects smaller or
ized applications like fluorescence, phase-contrast, confocal, thinner than 0.2 μm (such as a single ribosome or cytoplasmic
and polarizing microscopy, are all based on the interaction of microfilament) cannot be distinguished with this instrument.
light with tissue components and are used to reveal and study Likewise, two structures such as mitochondria will be seen as
tissue features. only one object if they are separated by less than 0.2 μm. The
microscope’s resolving power determines the quality of the
Bright-Field Microscopy image, its clarity and richness of detail, and depends mainly on
With the bright-field microscope stained tissue is examined the quality of its objective lens. Magnification is of value only
with ordinary light passing through the preparation. As shown when accompanied by high resolution. Objective lenses pro-
in Figure 1–3, the microscope includes an optical system and viding higher magnification are designed to also have higher
mechanisms to move and focus the specimen. The optical resolving power. The eyepiece lens only enlarges the image
components are the condenser focusing light on the object obtained by the objective and does not improve resolution.
to be studied; the objective lens enlarging and projecting the Virtual microscopy, typically used for study of bright-
image of the object toward the observer; and the eyepiece field microscopic preparations, involves the conversion of a
Light Microscopy 5
C H A P T E R
bright-field microscope. When certain cellular substances are irradiated by light of
a proper wavelength, they emit light with a longer wave-
Eyepiece Interpupillar
adjustment
length—a phenomenon called fluorescence. In fluores-
Binocular
tubes Head cence microscopy, tissue sections are usually irradiated with
ultraviolet (UV) light and the emission is in the visible portion
of the spectrum. The fluorescent substances appear bright on
Stand
a dark background. For fluorescent microscopy the instru-
1
Measuring
ment has a source of UV or other light and filters that select
N R
a b
Components of cells are often stained with compounds visible by filaments show nuclei with blue fluorescence and actin filaments
fluorescence microscopy. stained green. Important information such as the greater density
(a) Acridine orange binds nucleic acids and causes DNA in cell of microfilaments at the cell periphery is readily apparent. (Both
nuclei (N) to emit yellow light and the RNA-rich cytoplasm (R) to X500)
appear orange in these cells of a kidney tubule. (Figure 1–4b, used with permission from Drs Claire E. Walczak
and Rania Rizk, Indiana University School of Medicine,
(b) Cultured cells stained with DAPI (4′,6-diamino-2-phenylindole) Bloomington.)
that binds DNA and with fluorescein-phalloidin that binds actin
a b c
Living neural crest cells growing in culture appear differently in-phase light differently and produce an image of these features
with various techniques of light microscopy. Here the same field in all the cells.
of unstained cells, including two differentiating pigment cells, is (c) Differential interference microscopy: Cellular details are
shown using three different methods (all X200): highlighted in a different manner using Nomarski optics. Phase-
(a) Bright-field microscopy: Without fixation and staining, only contrast microscopy, with or without differential interference, is
the two pigment cells can be seen. widely used to observe live cells grown in tissue culture.
(b) Phase-contrast microscopy: Cell boundaries, nuclei, and (Used with permission from Dr Sherry Rogers, Department of Cell
cytoplasmic structures with different refractive indices affect Biology and Physiology, University of New Mexico, Albuquerque, NM.)
Light Microscopy 7
C H A P T E R
Laser Polarizing Microscopy
Polarizing microscopy allows the recognition of stained or
unstained structures made of highly organized subunits.
When normal light passes through a polarizing filter, it exits
1
Scanner
vibrating in only one direction. If a second filter is placed in
Lens
Other
Focal plane out-of-focus
Specimen planes
a feature of crystalline substances or substances containing The wavelength in an electron beam is much shorter than that
highly oriented molecules, such as cellulose, collagen, micro- of light, allowing a 1000-fold increase in resolution.
tubules, and actin filaments.
The utility of all light microscopic methods is greatly Transmission Electron Microscopy
extended through the use of digital cameras. Many features The transmission electron microscope (TEM) is an imag-
of digitized histological images can be analyzed quantitatively ing system that permits resolution around 3 nm. This high
using appropriate software. Such images can also be enhanced resolution allows isolated particles magnified as much as
to allow objects not directly visible through the eyepieces to be 400,000 times to be viewed in detail. Very thin (40-90 nm),
examined on a monitor. resin-embedded tissue sections are typically studied by TEM
at magnifications up to approximately 120,000 times.
›â•ºELECTRON MICROSCOPY Figure 1–8a indicates the components of a TEM and the
basic principles of its operation: a beam of electrons focused
Transmission and scanning electron microscopes are based on using electromagnetic “lenses” passes through the tissue sec-
the interaction of tissue components with beams of electrons. tion to produce an image with black, white, and intermediate
3 mm
Anode Anode
Copper grid
Condensor lens with three sections Lens
Specimen Column
Objective lens holder Lens
Column Scanner
Intermediate lens Electron detector
Image on viewing
screen Specimen
Electron detector
with CCD camera
Electron microscopes are large instruments generally housed in a In a TEM image areas of the specimen through which electrons
specialized EM facility. passed appear bright (electron lucent), while denser areas or
(a) Schematic view of the major components of a transmission elec- those that bind heavy metal ions during specimen preparation
tron microscope (TEM), which is configured rather like an upside- absorb or deflect electrons and appear darker (electron dense).
down light microscope. With the microscope column in a vacuum, a Such images are therefore always black, white, and shades of gray.
metallic (usually tungsten) filament (cathode) at the top emits elec- (b) The scanning electron microscope (SEM) has many similarities
trons that travel to an anode with an accelerating voltage between to a TEM. However, here the focused electron beam does not pass
60 and 120 kV. Electrons passing through a hole in the anode form a through the specimen, but rather is moved sequentially (scanned)
beam that is focused electromagnetically by circular electric coils from point to point across its surface similar to the way an electron
in a manner analogous to the effect of optical lenses on light. beam is scanned across a television tube or screen. For SEM speci-
The first lens is a condenser focusing the beam on the sec- mens are coated with metal atoms with which the electron beam
tion. Some electrons interact with atoms in the section, being interacts, producing reflected electrons and newly emitted secondary
absorbed or scattered to different extents, while others are simply electrons. All of these are captured by a detector and transmitted to
transmitted through the specimen with no interaction. Electrons amplifiers and processed to produce a black-and-white image on the
reaching the objective lens form an image that is then magnified monitor. The SEM shows only surface views of the coated specimen
and finally projected on a fluorescent screen or a charge-coupled but with a striking 3D, shadowed quality. The inside of organs or cells
device (CCD) monitor and camera. can be analyzed after sectioning to expose their internal surfaces.
Autoradiography 9
shades of gray regions. These regions of an electron micro- layer of heavy metal (often gold) which reflects electrons in
graph correspond to tissue areas through which electrons a beam scanning the specimen. The reflected electrons are
C H A P T E R
passed readily (appearing brighter or electron-lucent) and captured by a detector, producing signals that are processed
areas where electrons were absorbed or deflected (appearing to produce a black-and-white image. SEM images are usually
darker or more electron-dense). To improve contrast and reso- easy to interpret because they present a three-dimensional
lution in TEM, compounds with heavy metal ions are often view that appears to be illuminated in the same way that large
added to the fixative or dehydrating solutions used for tissue objects are seen with highlights and shadows caused by light.
preparation. These include osmium tetroxide, lead citrate,
1
and uranyl compounds, which bind cellular macromolecules,
›â•ºAUTORADIOGRAPHY
G
G
a b
Autoradiographs are tissue preparations in which particles called (a) Black grains of silver from the light-sensitive material coating
silver grains indicate the cells or regions of cells in which specific the specimen are visible over cell regions with secretory granules
macromolecules were synthesized just prior to fixation. Shown and the duct indicating glycoprotein locations. (X1500)
here are autoradiographs from the salivary gland of a mouse (b) The same tissue prepared for TEM autoradiography shows sil-
injected with 3H-fucose 8 hours before tissue fixation. Fucose was ver grains with a coiled or amorphous appearance again localized
incorporated into oligosaccharides, and the free 3H-fucose was mainly over the granules (G) and in the gland lumen (L). (X7500)
removed during fixation and sectioning of the gland. Autoradio- (Figure 1–9b, used with permission from Drs Ticiano G. Lima and
graphic processing and microscopy reveal locations of newly syn- A. Antonio Haddad, School of Medicine, Ribeirão Preto, Brazil.)
thesized glycoproteins containing that sugar.
10 CHAPTER 1â•… ■â•… Histology & Its Methods of Study
›â•ºCELL & TISSUE CULTURE activity. For enzyme histochemistry (1) tissue sections are
immersed in a solution containing the substrate of the enzyme
Live cells and tissues can be maintained and studied outside to be localized; (2) the enzyme is allowed to act on its sub-
the body in culture (in vitro). In the organism (in vivo), cells strate; (3) the section is then put in contact with a marker
are bathed in fluid derived from blood plasma and containing compound that reacts with a product of the enzymatic action
many different molecules required for survival and growth. on the substrate; and (4) the final product from the marker,
Cell culture allows the direct observation of cellular behavior which must be insoluble and visible by light or electron
under a phase-contrast microscope and many experiments microscopy, precipitates over the site of the enzymes, identify-
technically impossible to perform in the intact animal can be ing their location.
accomplished in vitro. Examples of enzymes that can be detected histochemi-
The cells and tissues are grown in complex solutions of cally include the following:
known composition (salts, amino acids, vitamins) to which ■⌀ Phosphatases, which remove phosphate groups from
serum or specific growth factors are added. Cells to be cultured macromolecules (Figure 1–10).
are dispersed mechanically or enzymatically from a tissue or ■⌀ Dehydrogenases, which transfer hydrogen ions from
organ and placed with sterile procedures in a clear dish to one substrate to another, such as many enzymes of the
which they adhere, usually as a single layer (Figure 1–5). Such citric acid (Krebs) cycle, allowing histochemical identifi-
preparations are called primary cell cultures. Some cells can cation of such enzymes in mitochondria.
be maintained in vitro for long periods because they become ■⌀ Peroxidase, which promotes the oxidation of sub-
immortalized and constitute a permanent cell line. Most cells strates with the transfer of hydrogen ions to hydrogen
obtained from normal tissues have a finite, genetically pro- peroxide.
grammed life span. However certain changes (some related
to oncogenes; see Chapter 3) can promote cell immortality, a
process called transformation, and are similar to the initial › ╺╺ MEDICAL APPLICATION
changes in a normal cell’s becoming a cancer cell. Improve-
Many enzyme histochemical procedures are used in the
ments in culture technology and use of specific growth factors
medical laboratory, including Perls’ Prussian blue reaction for
now allow most cell types to be maintained in vitro.
iron (used to diagnose the iron storage diseases, hemochro-
As shown in Chapter 2, incubation of living cells in vitro
matosis and hemosiderosis), the PAS-amylase and alcian blue
with a variety of new fluorescent compounds that are seques-
reactions for polysaccharides (to detect glycogenosis and
tered and metabolized in specific compartments of the cell
mucopolysaccharidosis), and reactions for lipids and sphin-
provides a new approach to understanding these compart-
golipids (to detect sphingolipidosis).
ments both structurally and physiologically. Other histologic
techniques applied to cultured cells have been particularly
important for understanding the locations and functions of
microtubules, microfilaments, and other components of the
cytoskeleton. ›â•ºVISUALIZING SPECIFIC MOLECULES
A specific macromolecule present in a tissue section may also
be identified by using tagged compounds or macromolecules
› ╺╺ MEDICAL APPLICATION that bind specifically with the molecule of interest. The com-
Cell culture is very widely used to study molecular changes pounds that interact with the molecule must be visible with
that occur in cancer; to analyze infectious viruses, myco- the light or electron microscope, often by being tagged with a
plasma, and some protozoa; and for many routine genetic or detectible label. The most commonly used labels are fluores-
chromosomal analyses. Cervical cancer cells from a patient cent compounds, radioactive atoms that can be detected with
later identified as Henrietta Lacks, who died from the disease autoradiography, molecules of peroxidase or other enzymes
in 1951, were used to establish one of the first cell lines, that can be detected with histochemistry, and metal (usually
called HeLa cells, which are still used in research on cellular gold) particles that can be seen with light and electron micros-
structure and function throughout the world. copy. These methods can be used to detect and localize specific
sugars, proteins, and nucleic acids.
Visualizing Specific Molecules 11
C H A P T E R
■⌀ Phalloidin, a compound extracted from mushroom,
Amanita phalloides, interacts strongly with the actin pro-
tein of microfilaments.
L ■⌀ Protein A, purified from Staphylococcus aureus bacte-
ria, binds to the Fc region of antibody molecules, and
can therefore be used to localize naturally occurring or
1
applied antibodies bound to cell structures.
Histology & Its Methods of Study ╇ ■╇ Visualizing Specific Molecules
■⌀ Lectins, glycoproteins derived mainly from plant seeds,
bind to carbohydrates with high affinity and specificity.
Different lectins bind to specific sugars or sequences of
sugar residues, allowing fluorescently labeled lectins to
be used to stain specific glycoproteins or other macro-
molecules bearing specific sequences of sugar residues.
Immunohistochemistry
A highly specific interaction between macromolecules is that
between an antigen and its antibody. For this reason labeled
antibodies are routinely used in immunohistochemistry
L L
to identify and localize many specific proteins, not just
those with enzymatic activity that can be demonstrated by
histochemistry.
aa The body’s immune cells interact with and produce anti-
bodies against other macromolecules—called antigens—that
are recognized as “foreign,” not a normal part of the organism,
and potentially dangerous. Antibodies belong to the immu-
noglobulin family of glycoproteins and are secreted by lym-
phocytes. These molecules normally bind specifically to their
Ly provoking antigens and help eliminate them.
Widely applied for both research and diagnostic pur-
poses, every immunohistochemical technique requires an
antibody against the protein that is to be detected. This means
Ly that the protein must have been previously purified using bio-
chemical or molecular methods so that antibodies against it
can be produced. To produce antibodies against protein x of a
certain animal species (eg, a human or rat), the isolated pro-
tein is injected into an animal of another species (eg, a rabbit
or a goat). If the protein’s amino acid sequence is sufficiently
b N different for this animal to recognize it as foreign—that is, as
an antigen—the animal will produce antibodies against the
protein.
(a) Micrograph of cross sections of kidney tubules treated Different groups (clones) of lymphocytes in the injected
histochemically to demonstrate alkaline phosphatases (with animal recognize different parts of protein x and each clone
maximum activity at an alkaline pH) showing strong activity of produces an antibody against that part. These antibodies are
this enzyme at the apical surfaces of the cells at the lumens (L)
of the tubules. (X200)
collected from the animal’s plasma and constitute a mixture
of polyclonal antibodies, each capable of binding a different
(b) TEM image of a kidney cell in which acid phosphatase has
been localized histochemically in three lysosomes (Ly) near the region of protein x.
nucleus (N). The dark material within these structures is lead It is also possible, however, to inject protein x into a
phosphate that precipitated in places with acid phosphatase mouse and a few days later isolate the activated lymphocytes
activity. (X25,000) and place them into culture. Growth and activity of these cells
(Figure 1–10b, used with permission from Dr Eduardo
can be prolonged indefinitely by fusing them with lymphocytic
Katchburian, Department of Morphology, Federal University of
São Paulo, Brazil.) tumor cells to produce hybridoma cells. Different hybridoma
clones produce different antibodies against the several parts
12 CHAPTER 1â•… ■â•… Histology & Its Methods of Study
of protein x and each clone can be isolated and cultured sepa- The indirect method is used more widely in research and
rately so that the different antibodies against protein x can be pathologic tests because it is more sensitive, with the extra
collected separately. Each of these antibodies is a monoclo- level of antibody binding serving to amplify the visible signal.
nal antibody. An advantage to using a monoclonal antibody Moreover, the same preparation of labeled secondary antibody
rather than polyclonal antibodies is that it can be selected to can be used in studies with different primary antibodies (spe-
be highly specific and to bind strongly to the protein to be cific for different antigens) as long as all these are made in the
detected, with less nonspecific binding to other proteins that same species. There are other indirect methods that involve the
are similar to the one of interest. use of other intermediate molecules, such as the biotin-avidin
In immunohistochemistry a tissue section that one technique, which are also used to amplify detection signals.
believes contains the protein of interest is incubated in a solu- Examples of indirect immunocytochemistry are shown in
tion containing antibody (either monoclonal or polyclonal) Figure 1–12, demonstrating the use of this method with cells
against this protein. The antibody binds specifically to the in culture or after tissue sectioning for both light microscopy
protein and after a rinse the protein’s location in the tissue or and TEM.
cells can be seen with either the light or electron microscope
by visualizing the antibody. Antibodies are commonly tagged › ╺╺ MEDICAL APPLICATION
with fluorescent compounds, with peroxidase or alkaline
Because cells in some diseases, including many cancer cells,
phosphatase for histochemical detection, or with electron-
often produce proteins unique to their pathologic condition,
dense gold particles for TEM.
immunohistochemistry can be used by pathologists to diag-
As Figure 1–11 indicates, there are direct and indirect
nose many diseases, including certain types of tumors and
methods of immunocytochemistry. The direct method just
some virus-infected cells. Table 1-1 shows some applications
involves a labeled antibody that binds the protein of interest.
of immunocytochemistry routinely used in clinical practice.
Indirect immunohistochemistry involves sequential
application of two antibodies and additional washing steps. The
(primary) antibody specifically binding the protein of interest Hybridization Techniques
is not labeled. The detectible tag is conjugated to a second- Hybridization usually implies the specific binding between
ary antibody made in an animal species different (“foreign”) two single strands of nucleic acid, which occurs under appro-
from that which made the primary antibody. For example, pri- priate conditions if the strands are complementary. The greater
mary antibodies made by mouse lymphocytes (such as most the similarities of their nucleotide sequences, the more read-
monoclonal antibodies) are specifically recognized and bound ily the complementary strands form “hybrid” double-strand
by antibodies made in a rabbit or goat injected with mouse molecules. Hybridization at stringent conditions allows the
antibody immunoglobulin. specific identification of sequences in genes or RNA. This can
Labeled
secondary
Labeled Unlabeled antibody
antibody primary
antibody
Antigen Antigen
Tissue section
Glass slide
Direct Indirect
Immunocytochemistry (or immunohistochemistry) can be direct labeled secondary antibody is obtained that was (1) made in
or indirect. Direct immunocytochemistry (left) uses an antibody another species against immunoglobulin proteins (antibodies)
made against the tissue protein of interest and tagged directly from the species in which the primary antibodies were made and
with a label such as a fluorescent compound or peroxidase. When (2) labeled with a fluorescent compound or peroxidase. When
placed with the tissue section on a slide, these labeled antibod- the labeled secondary antibody is applied to the tissue section, it
ies bind specifically to the protein (antigen) against which they specifically binds the primary antibodies, indirectly labeling the
were produced and can be visualized by the appropriate method. protein of interest on the slide. Because more than one labeled
Indirect immunocytochemistry (right) uses first a primary secondary antibody can bind each primary antibody molecule,
antibody made against the protein (antigen) of interest and labeling of the protein of interest is amplified by the indirect
applied to the tissue section to bind its specific antigen. Then a method.
Visualizing Specific Molecules 13
1 C H A P T E R
Histology & Its Methods of Study ╇ ■╇ Visualizing Specific Molecules
a c
╇
TABLE 1-1 Examples of specific antigens with diagnostic importance.
Antigens Diagnosis
occur with cellular DNA or RNA when nucleic acid sequences › ╺╺ MEDICAL APPLICATION
in solution are applied directly to prepared cells and tissue sec-
Warts on the skin of the genitals and elsewhere are due to
tions, a procedure called in situ hybridization (ISH).
infection with the human papilloma virus (HPV) which causes
This technique is ideal for (1) determining if a cell has
the characteristic benign proliferative growth. As shown in
a specific sequence of DNA, such as a gene or part of a gene
Figure 1–12 such virus-infected cells can often be demon-
(Figure 1–13), (2) identifying the cells containing specific
strated by ISH. Certain cancer cells with unique or elevated
messenger RNAs (mRNAs) (in which the corresponding
expression of specific genes are also localized in tumors and
gene is being transcribed), or (3) determining the localiza-
studied microscopically by ISH.
tion of a gene in a specific chromosome. DNA and RNA of
the cells must be initially denatured by heat or other agents
to become completely single-stranded and the nucleotide
sequences of interest are detected with probes consisting of
single-stranded complementary DNA (cDNA). The probe
›â•ºINTERPRETATION OF STRUCTURES
IN TISSUE SECTIONS
may be obtained by cloning, by polymerase chain reaction
(PCR) amplification of the target sequence, or by chemical In studying and interpreting stained tissue sections, it is
synthesis if the desired sequence is short. The probe is tagged important to remember that microscopic preparations are the
with nucleotides containing a radioactive isotope (localized by end result of a series of processes that began with collecting
autoradiography) or modified with a small compound such as the tissue and ended with mounting a coverslip on the slide.
digoxigenin (identified by immunocytochemistry). A solution Certain steps in this procedure may distort the tissues slightly,
containing the probe is placed over the specimen under con- producing minor structural abnormalities called artifacts not
ditions allowing hybridization and after the excess unbound present in the living tissue.
probe is washed off, the localization of the hybridized probe is One such distortion is minor shrinkage of cells or tissue
revealed through its label. regions produced by the fixative, by the ethanol, or by the heat
needed for paraffin embedding. Shrinkage can create artificial
spaces between cells and other tissue components. Such spaces
FIGURE 1–13╇ In situ hybridization (ISH). can also result from the loss of lipids or low-molecular-weight
substances not preserved by the fixative or removed by the
dehydrating and clearing fluids. Slight cracks in sections may
also appear as large spaces in the tissue.
Other artifacts may include small wrinkles in the section
(which the novice may confuse with linear structures in tissue)
and precipitates from the stain (which may be confused with
cellular structures such as cytoplasmic granules). Students must
be aware of the existence of artifacts and able to recognize them.
Another difficulty in the study of histologic sections is the
impossibility of differentially staining all tissue components on
one slide. A single stain can seldom demonstrate well nuclei,
mitochondria, lysosomes, basement membranes, elastic fibers,
etc. With the light microscope, it is necessary to examine prepa-
rations stained by different methods before an idea of the whole
composition and structure of a cell or tissue can be obtained. The
TEM allows the observation of cells with all its internal struc-
tures and surrounding ECM components, but only a few cells in
a tissue can be conveniently studied in these very small samples.
Finally, when a structure’s three-dimensional volume is
cut into very thin sections, the sections appear microscopically
to have only two dimensions: length and width. When examin-
In situ hybridization of this tissue section with probes for the ing a section under the microscope, the viewer must always keep
human papilloma virus (HPV) reveals the presence of many
cells containing the virus. The section was incubated with a
in mind that components are missing in front of and behind
solution containing a digoxigenin-labeled complementary what is being seen because many tissue structures are thicker
DNA (cDNA) probe for the HPV DNA. The probe was then than the section. Round structures seen microscopically may
visualized by direct immunohistochemistry using peroxidase- actually be portions of spheres or tubes. Because structures in
labeled antibodies against digoxigenin. This procedure stains a tissue have different orientations, their two-dimensional (2D)
brown only those cells containing HPV. (X400; H&E)
(Used with permission from Dr Jose E. Levi, Virology Lab,
appearance will also vary depending on the plane of section. A
Institute of Tropical Medicine, University of São Paulo, Brazil.) single convoluted tube will appear in a tissue section as many
separate rounded or oval structures (Figure 1–14).
Interpretation of Structures in Tissue Sections 15
1 C H A P T E R
Histology & Its Methods of Study ╇ ■╇ Interpretation of Structures in Tissue Sections
In thin sections 3D structures appear to have only two dimensions.
Such images must be interpreted correctly to understand the actual
structure of tissue and organ components. For example, blood ves-
sels and other tubular structures appear in sections as round or oval
shapes whose size and shape depend on the transverse or oblique
angle of the cut. A highly coiled tube will appear as several round
and oval structures. In TEM sections of cells, round structures may
represent spherical organelles or transverse cuts through tubular
organelles such as mitochondria. It is important to develop such
interpretive skill to understand tissue and cell morphology in micro-
scopic preparations.
■⌀ Indirect immunohistochemistry uses an unlabeled primary anti- Interpretation of Structures in Tissue Sections
body that is detected bound to its antigen with labeled secondary ■⌀ Many steps in tissue processing, slide preparation, and staining can
antibodies. introduce minor artifacts such as spaces and precipitates that are
■⌀ The indirect immunohistochemical method is more commonly not normally present in the living tissue and must be recognized.
used because the added level of antibody binding amplifies the sig- ■⌀ Sections of cells or tissues are essentially 2D planes through 3D
nal detected and provides greater technical flexibility. structures, and understanding this fact is important for their cor-
■⌀ Specific gene sequences or mRNAs of cells can be detected micro- rect interpretation and study.
scopically using labeled complementary DNA (cDNA) probes in a
technique called in situ hybridization (ISH).
1. In preparing tissue for routine light microscopic study, which 7. Microscopic autoradiography uses radioactivity and can be employed
procedure immediately precedes clearing the specimen with an to study what features in a tissue section?
organic solvent? a. The types of enzymes found in various cell locations
a. Dehydration b. Cellular sites where various macromolecules are synthesized
b. Fixation c. The sequences of mRNA made in the cells
c. Staining d. The dimensions of structures within the cells
d. Clearing e. The locations of genes transcribed for specific mRNA
e. Embedding
8. To identify and localize a specific protein within cells or the extracel-
2. Which of the following staining procedures relies on the cationic lular matrix one would best use what approach?
and anionic properties of the material to be stained? a. Autoradiography
a. Enzyme histochemistry b. Enzyme histochemistry
b. Periodic acid-Schiff reaction c. Immunohistochemistry
c. Hematoxylin & eosin staining d. Transmission electron microscopy
d. Immunohistochemistry e. Polarizing microscopy
e. Metal impregnation techniques
9. In situ hybridization is a histological technique used to visualize what
3. In a light microscope used for histology, resolution and magnifica- type of macromolecule?
tion of cells are largely dependent on which component? a. Proteins
a. Condenser b. Carbohydrates
b. Objective lens c. Certain enzymes
c. Eyepieces or ocular lenses d. Nucleic acids
d. Specimen slide e. Lipids
e. The control for illumination intensity
10. Hospital laboratories frequently use unfixed, frozen tissue specimens
4. Cellular storage deposits of glycogen, a free polysaccharide, could sectioned with a cryostat for rapid staining, microscopic examina-
best be detected histologically using what procedure? tion, and diagnosis of pathological conditions. Besides saving much
a. Autoradiography time by avoiding fixation and procedures required for paraffin
b. Electron microscopy embedding, frozen sections retain and allow study of what macro-
c. Enzyme histochemistry molecules normally lost in the paraffin procedure?
d. Hematoxylin & eosin staining a. Carbohydrates
e. Periodic acid-Schiff reaction b. Small mRNA
c. Basic proteins
5. Adding heavy metal compounds to the fixative and ultrathin sec- d. Acidic proteins
tioning of the embedded tissue with a glass knife are techniques e. Lipids
used for which histological procedure?
a. Scanning electron microscopy
b. Fluorescent microscopy
c. Enzyme histochemistry
d. Confocal microscopy
e. Transmission electron microscopy
6. Resolution in electron microscopy greatly exceeds that of light
microscopy due to which of the following?
a. The wavelength of the electrons in the microscope beam is
shorter than that of a beam of light.
b. The lenses of an electron microscope are of greatly improved
quality.
c. For electron microscopy the tissue specimen does not require
staining.
d. The electron microscope allows much greater magnification of
a projected image than a light microscope provides.
e. An electron microscope can be much more finely controlled
than a light microscope.
Answers: 1a, 2c, 3b, 4e, 5e, 6a, 7b, 8c, 9d, 10e
C H A P T E R
CELL DIFFERENTIATION
2 The Cytoplasm
17 Proteasomes
Mitochondria
37
38
THE PLASMA MEMBRANE 17
Peroxisomes 39
Transmembrane Proteins & Membrane Transport 19
Vesicular Transport: Endocytosis & Exocytosis 21 THE CYTOSKELETON 42
Signal Reception & Transduction 26 Microtubules 43
CYTOPLASMIC ORGANELLES 27 Microfilaments (Actin Filaments) 44
Intermediate Filaments 45
Ribosomes 27
Endoplasmic Reticulum 28 INCLUSIONS 47
Golgi Apparatus 31 SUMMARY OF KEY POINTS 51
Secretory Granules 33
ASSESS YOUR KNOWLEDGE 52
Lysosomes 34
17
18 CHAPTER 2â•… ■â•… The Cytoplasm
C H A P T E R
Polar head group Nonpolar fatty acid chains
(hydrophilic) (hydrophobic)
O
Saturated CH3
CH2 O C
fatty acid
O
2
(straight) CH3
CH O C
Phospholipids
Hydrophilic surface
Hydrophobic region
Extracellular fluid
Hydrophilic surface
Cholesterol
Cytoplasm
(a) Membranes of animal cells have as their major lipid com- throughout the lipid bilayer; cholesterol affects the packing of the
ponents phospholipids and cholesterol. A phospholipid is fatty acid chains, with a major effect on membrane fluidity. The
amphipathic, with a phosphate group charge on the polar head outer layer of the cell membrane also contains glycolipids with
and two long, nonpolar fatty acid chains, which can be straight extended carbohydrate chains.
(saturated) or kinked (at an unsaturated bond). Membrane choles- Sectioned, osmium-fixed cell membrane may have a faint trilam-
terol is present in about the same amount as phospholipid. inar appearance with the transmission electron microscope (TEM),
(b) The amphipathic nature of phospholipids produces the bilayer showing two dark (electron-dense) lines enclosing a clear (electron-
structure of membranes as the charged (hydrophilic) polar heads lucent) band. Reduced osmium is deposited on the hydrophilic
spontaneously form each membrane surface, in direct contact phosphate groups present on each side of the internal region of
with water, and the hydrophobic nonpolar fatty acid chains are fatty acid chains where osmium is not deposited. The “fuzzy” mate-
buried in the membrane’s middle, away from water. Cholesterol rial on the outer surface of the membrane represents the glycocalyx
molecules are also amphipathic and are interspersed less evenly of oligosaccharides of glycolipids and glycoproteins. (X100,000)
cholesterol and saturated fatty acids which reduce lipid fluid- molecules cross the membrane by the general mechanisms
ity. This together with the presence of scaffold proteins that shown schematically in Figure 2–5 and explained as follows:
maintain spatial relationships between enzymes and signal-
ing proteins allows the proteins assembled within lipid rafts to
■⌀ Diffusion transports small, nonpolar molecules directly
through the lipid bilayer. Lipophilic (fat-soluble) mol-
remain in close proximity and interact more efficiently. ecules diffuse through membranes readily, water very
slowly.
Transmembrane Proteins & MembraneTransport ■⌀ Channels are multipass proteins forming transmembrane
The plasma membrane is the site where materials are pores through which ions or small molecules pass selec-
exchanged between the cell and its environment. Most small tively. Cells open and close specific channels for Na+,
20 CHAPTER 2â•… ■â•… The Cytoplasm
2 E face
Peripheral protein 1
Transmembrane protein
Lipid
P face
(a) The fluid mosaic model of membrane structure emphasizes center. Splitting occurs along the line of weakness formed by
that the phospholipid bilayer of a membrane also contains pro- the fatty acid tails of phospholipids. Electron microscopy of such
teins inserted in it or associated with its surface (peripheral pro- cryofracture preparation replicas provides a useful method for
teins) and that many of these proteins move within the fluid lipid studying membrane structures. Most of the protruding mem-
phase. Integral proteins are firmly embedded in the lipid layers; brane particles seen (1) are proteins or aggregates of proteins
those that completely span the bilayer are called transmembrane that remain attached to the half of the membrane adjacent to
proteins. Hydrophobic amino acids of these proteins interact with the cytoplasm (P or protoplasmic face). Fewer particles are found
the hydrophobic fatty acid portions of the membrane lipids. Both attached to the outer half of the membrane (E or extracellular
the proteins and lipids may have externally exposed oligosaccha- face). Each protein bulging on one surface has a corresponding
ride chains. depression (2) on the opposite surface.
(b) When cells are frozen and fractured (cryofracture), the lipid
bilayer of membranes is often cleaved along the hydrophobic
K+, Ca2+ and other ions in response to various physio- down a concentration gradient due to its kinetic energy. In
logical stimuli. Water molecules usually cross the plasma contrast, membrane pumps are enzymes engaged in active
membrane through channel proteins called aquaporins. transport, utilizing energy from the hydrolysis of adenos-
■⌀ Carriers are transmembrane proteins that bind small ine triphosphate (ATP) to move ions and other solutes across
molecules and translocate them across the membrane via membranes, against often steep concentration gradients.
conformational changes. Because they consume ATP pumps they are often referred to
as ATPases.
Diffusion, channels, and carrier proteins operate pas- These transport mechanisms are summarized with addi-
sively, allowing movement of substances across membranes tional detail in Table 2–2.
The Plasma Membrane 21
C H A P T E R
Interstitial fluid
Phospholipid
2
Glycolipid Carbohydrate
Glycoprotein
Nonpolar tails
of phospholipid Cholesterol Protein
molecule
Integral protein
Peripheral protein
Filaments of
cytoskeleton
Cytosol
1. Physical barrier: Establishes a flexible boundary, protects cellular contents, 3. Electrochemical gradients: Establishes and maintains an electrical
and supports cell structure. Phospholipid bilayer separates substances charge difference across the plasma membrane.
inside and outside the cell. 4. Communication: Contains receptors that recognize and respond to
2. Selective permeability: Regulates entry and exit of ions, nutrients, molecular signals.
and waste molecules through the membrane.
Both protein and lipid components often have covalently connections, and as selective gateways for molecules entering
attached oligosaccharide chains exposed at the external mem- the cell.
brane surface. These contribute to the cell’s glycocalyx, which Transmembrane proteins often have multiple hydrophobic
provides important antigenic and functional properties to the regions buried within the lipid bilayer to produce a channel or
cell surface. Membrane proteins serve as receptors for vari- other active site for specific transfer of substances through the
ous signals coming from outside cells, as parts of intercellular membrane.
Vesicular â•›Transport: â•›Endocytosis & â•›Exocytosis which project from the cell in a process dependent on
cytoskeletal changes. Fusion of the membranous folds
Macromolecules normally enter cells by being enclosed within
encloses the bacterium in an intracellular vacuole called
folds of plasma membrane (often after binding specific mem-
a phagosome, which then merges with a lysosome
brane receptors) which fuse and pinch off internally as cyto-
for degradation of its contents as discussed later in this
plasmic vesicles (or vacuoles) in a general process known as
chapter.
endocytosis. Three major types of endocytosis are recog-
nized, as summarized in Table 2–2 and Figure 2–6. 2. Pinocytosis (“cell drinking”) involves smaller invagina-
tions of the cell membrane which fuse and entrap extra-
1. Phagocytosis (“cell eating”) is the ingestion of particles cellular fluid and its dissolved contents. The resulting
such as bacteria or dead cell remnants. Certain blood- pinocytotic vesicles (~80 nm in diameter) then pinch
derived cells, such as macrophages and neutrophils, are off inwardly from the cell surface and either fuse with
specialized for this activity. When a bacterium becomes lysosomes or move to the opposite cell surface where
bound to the surface of a neutrophil, it becomes sur- they fuse with the membrane and release their contents
rounded by extensions of plasmalemma and cytoplasm outside the cell. The latter process, called transcytosis,
22 CHAPTER 2â•… ■â•… The Cytoplasm
2 C H A P T E R
The Cytoplasm╇ ■╇ The Plasma Membrane
(a) Simple diffusion (b) Channel (c) Carrier/pump
Lipophilic and some small, uncharged molecules can cross mem- conformations and release the molecule to the other side of the
branes by simple diffusion (a). membrane.
Most ions cross membranes in multipass proteins called channels Diffusion, channels and most carrier proteins translocate sub-
(b) whose structures include transmembrane ion-specific pores. stances across membranes using only kinetic energy. In contrast,
Many other larger, water-soluble molecules require binding pumps are carrier proteins for active transport of ions or other
to sites on selective carrier proteins (c), which then change their solutes and require energy derived from ATP.
Exocytosis of macromolecules made by cells occurs via are called exosomes, which can fuse with other cells transfer-
either of two pathways: ring their contents and membranes.
■⌀ Constitutive secretion is used for products that are
released from cells continuously, as soon as synthesis is Signal Reception & Transduction
complete, such as collagen subunits for the ECM. Cells in a multicellular organism communicate with one
■⌀ Regulated secretion occurs in response to signals com- another to regulate tissue and organ development, to control
ing to the cells, such as the release of digestive enzymes their growth and division, and to coordinate their functions.
from pancreatic cells in response to specific stimuli. Many adjacent cells form communicating gap junctions that
Regulated exocytosis of stored products from epithelial couple the cells and allow exchange of ions and small mol-
cells usually occurs specifically at the apical domains of ecules (see Chapter 4).
cells, constituting a major mechanism of glandular secretion Cells also use about 25 families of receptors to detect
(see Chapter 4). and respond to various extracellular molecules and physical
stimuli. Each cell type in the body contains a distinctive set
Portions of the cell membrane become part of the endo-
of cell surface and cytoplasmic receptor proteins that enable
cytotic vesicles or vacuoles during endocytosis; during exo-
it to respond to a complementary set of signaling molecules
cytosis, membrane is returned to the cell surface. This process
in a specific, programmed way. Cells bearing receptors for a
of membrane movement and recycling is called membrane
specific ligand are referred to as target cells for that molecule.
trafficking (Figure 2–7a). Trafficking of membrane com-
The routes of signal molecules from source to target provide
ponents occurs continuously in most cells and is not only
one way to categorize the signaling process:
crucial for maintaining the cell but also for physiologically
important processes such as reducing blood lipid levels. ■⌀ In endocrine signaling, the signal molecules (here
In many cells subpopulations of vacuoles and tubules called hormones) are carried in the blood from their
within the endosomal compartment accumulate small vesicles sources to target cells throughout the body.
within their lumens by further invaginations of their limiting ■⌀ In paracrine signaling, the chemical ligand diffuses in
membranes, becoming multivesicular bodies. While multi- extracellular fluid but is rapidly metabolized so that its
vesicular bodies may merge with lysosomes for selective deg- effect is only local on target cells near its source.
radation of their content, this organelle may also fuse with the ■⌀ In synaptic signaling, a special kind of paracrine inter-
plasma membrane and release the intralumenal vesicles out- action, neurotransmitters act on adjacent cells through
side the cell. The small (<120 nm diameter) vesicles released special contact areas called synapses (see Chapter 9).
24 CHAPTER 2â•… ■â•… The Cytoplasm
╇
TABLE 2–2 Mechanisms of transport across the plasma membrane.
Process Type of Movement Example
PASSIVE PROCESSES Movement of substances down a concentration gradient due to the kinetic energy of the substance;
no expenditure of cellular energy is required; continues until equilibrium is reached (if unopposed)
Simple diffusion Unassisted net movement of small, nonpolar Exchange of oxygen and carbon dioxide
substances down their concentration gradient across a between blood and body tissues
selectively permeable membrane
Facilitated diffusion Movement of ions and small, polar molecules
down their concentration gradient; assisted across
a selectively permeable membrane by a transport
protein
â•…Channel-mediated Movement of ion down its concentration gradient Na+ moves through Na+ channel into cell
through a protein channel
â•…Carrier-mediated Movement of small, polar molecule down its Transport of glucose into cells by glucose
concentration gradient by a carrier protein carrier
Osmosis Diffusion of water across a selectively permeable Solutes in blood in systemic capillaries
membrane; direction is determined by relative solute “pulls” fluid from interstitial space back into
concentrations; continues until equilibrium the blood
is reached
ACTIVE PROCESSES Movement of substances requires expenditure of cellular energy
Active transport Transport of ions or small molecules across the
membrane against a concentration gradient by
transmembrane protein pumps
â•…Primary Movement of substance up its concentration gradient; Ca2+ pumps transport Ca2+ out of the cell
powered directly by ATP Na+/K+ pump moves Na+ out of cell and K+
into cell
â•…Secondary Movement of a substance up its concentration
gradient is powered by harnessing the movement of
a second substance (eg, Na+) down its concentration
gradient
â•…Symport Movement of substance up its concentration gradient Na+/glucose transport
in the same direction as Na+
â•…Antiport Movement of substance up its concentration gradient Na+/H+ transport
in the opposite direction from Na+
Vesicular transport Vesicle formed or lost as material is brought into a cell
or released from a cell
â•…Exocytosis Bulk movement of substance out of the cell by fusion Release of neurotransmitter by nerve cells
of secretory vesicles with the plasma membrane
â•…Endocytosis Bulk movement of substances into the cell by vesicles
forming at the plasma membrane
â•…Phagocytosis Type of endocytosis in which vesicles are formed as White blood cell engulfing a bacterium
particulate materials external to the cell are engulfed
by pseudopodia
â•…Pinocytosis Type of endocytosis in which vesicles are formed as Formation of small vesicles in capillary wall
interstital fluid is taken up by the cell to move substances
â•…Receptor-mediated endocytosis Type of endocytosis in which plasma membrane Uptake of cholesterol into cells
receptors first bind specific substances; receptor and
bound substance then taken up by the cell
The Plasma Membrane 25
C H A P T E R
Extracellular fluid
Pseudopodia
Particle
2
Plasma
Plasma Vesicle membrane
Cytoplasm
a Phagocytosis
b Pinocytosis
Receptors
Receptor
Ligand ligand Clathrin
complexes coat
Receptors
Coated pit
Apical
domain Dynamin
of cell Adaptor
membrane protein Clathrin
Coat
proteins Coated CP CP
recycled vesicle
CV
Receptor
recycling
Early
endosome
Late
endosome
Transcytosis b
Lysosomal
degradation
Basolateral domain
of cell membrane
Major steps during and after endocytosis are indicated diagram- ■⌀ Receptors and ligands may be carried to late endosomes and
matically in part a. Ligands bind with high affinity to specific then to lysosomes for degradation.
surface receptors, which then associate with specific cytoplasmic ■⌀ Ligands may be released from the receptors and the empty
proteins, including clathrin and adaptor proteins, and aggregate receptors sequestered into recycling endosomes and
in membrane regions to form coated pits. Clathrin facilitates returned to the cell surface for reuse.
invagination of the pits, and another peripheral membrane pro- ■⌀ Other endosomal vesicles containing ligands may move
tein, dynamin, forms constricting loops around the developing to and fuse with another cell surface, where the ligands
neck of the pit, causing the invagination to pinch off as a coated are released again outside the cell in the process of
vesicle. The clathrin lattice of coated pits (CP) and vesicles (CV) is transcytosis.
shown ultrastructurally in part b.
(Figure 2–7b, used with permission from Dr John Heuser,
Internalized vesicles lose their clathrin coats, which are
Department of Cell Biology and Physiology, Washington University
recycled, and fuse with other endosomes that comprise the endo-
School of Medicine, St. Louis, MO.)
somal compartment. Ligands may have different fates within the
endosomal compartment:
monophosphate (cAMP). Other second messengers include carrier proteins in the plasma for transport through the body.
1,2-diacylglycerol (DAG), and inositol 1,4,5-triphosphate (IP3). Such hormones are lipophilic and pass by diffusion through
The ionic changes or second messengers amplify the first signal cell membranes, binding to specific cytoplasmic receptor
and trigger a cascade of enzymatic activity, usually including proteins in target cells. With many steroid hormones, recep-
kinases, leading to changes in gene expression or cell behavior. tor binding activates that receptor, enabling the complex to
Second messengers may diffuse through the cytoplasm or be move into the nucleus and bind with high affinity to specific
retained locally by scaffold proteins for more focused amplifica- DNA sequences. This generally increases the level of tran-
tion of activity. scription of those genes. Each steroid hormone is recognized
Low molecular weight hydrophobic signaling molecules, by a different member of a family of homologous receptor
such as steroids and thyroid hormones, bind reversibly to proteins.
Cytoplasmic Organelles 27
C H A P T E R
Channel open
Ions
Ligand g
Ligand
2
Channel
1 A ligand binds to a
receptor, causing a Ions
conformational
connfor
format
ma ion
mational
al cha
change
nge
Ligand to
o activate
ac
activa receptor.
tivate recep
cep
ceptor
e tor
tor. Effector protein (eg, ion channel)
Protein and most small ligands are hydrophilic molecules that bind phosphorylate (and usually activate) other proteins upon ligand
transmembrane protein receptors to initiate changes in the target cell. binding. (c) G-protein–coupled receptors bind ligand, changing
(a) Channel-linked receptors bind ligands such as neurotrans- the conformation of its G-protein subunit, allowing it to bind GTP,
mitters and open to allow influx of specific ions. (b) Enzymatic and activating and releasing this protein to in turn activate other
receptors are usually protein kinases that are activated to proteins such as ion channels and adenyl cyclase.
ribosomal subunit is a highly folded ribosomal RNA (rRNA) Proper folding of new proteins is guided by protein chaper-
chain associated with more than 30 unique proteins; the core ones. Denatured proteins or those that cannot be refolded prop-
of the large subunit has three other rRNA molecules and erly are conjugated to the protein ubiquitin that targets them for
nearly 50 other basic proteins. breakdown by proteasomes (discussed below). As indicated in
The rRNA molecules in the ribosomal subunits not only Figure 2–9, proteins synthesized for use within the cytosol (eg,
provide structural support but also position transfer RNAs glycolytic enzymes) or for import into the nucleus and certain
(tRNA) molecules bearing amino acids in the correct “reading other organelles are synthesized on polyribosomes existing as
frame” and catalyze the formation of the peptide bonds. The isolated cytoplasmic clusters. Polyribosomes attached to mem-
more peripheral proteins of the ribosome seem to function branes of the endoplasmic reticulum (ER) translate mRNAs
primarily to stabilize the catalytic RNA core. coding for membrane proteins of the ER, the Golgi apparatus,
These ribosomal proteins are themselves synthesized in or the cell membrane; enzymes to be stored in lysosomes; and
cytoplasmic ribosomes, but are then imported to the nucleus proteins to undergo exocytosis from secretory vesicles.
where they associate with newly synthesized rRNA. The ribo-
somal subunits thus formed then move from the nucleus to
the cytoplasm where they are reused many times, for transla- Endoplasmic Reticulum
tion of any mRNA strand. The cytoplasm of most cells contains a convoluted membranous
During protein synthesis many ribosomes typically bind network called the endoplasmic reticulum (ER). As shown in
the same strand of mRNA to form larger complexes called Figure 2–10 this network (reticulum) extends from the surface
polyribosomes, or polysomes (Figure 2–9). In stained of the nucleus throughout most of the cytoplasm and encloses
preparations of cells polyribosomes are intensely basophilic a series of intercommunicating channels called cisternae
because of the numerous phosphate groups of the constitu- (L. cisternae, reservoirs). With a membrane surface up to 30 times
ent RNA molecules that act as polyanions. Thus, cytoplasmic that of the plasma membrane, the ER is a major site for vital
regions that stain intensely with hematoxylin and basic dyes, cellular activities, including biosynthesis of proteins and lipids.
such as methylene and toluidine blue, indicate sites of active Numerous polyribosomes attached to the membrane in some
protein synthesis. regions of ER allow two types of ER to be distinguished.
3′
FREE POLYRIBOSOMES 5′ ER-BOUND POLYRIBOSOMES 3′
5′
mRNA
Ribosome
Cisterna of
rough ER
Misfolded &
Proteins of cytosol denatured proteins
& cytoskeleton
Conjugated to ubiquitin Golgi apparatus processing & sorting
Free polyribosomes (not attached to the endoplasmic reticulum, polysomes attached to the membranes of ER. The proteins pro-
or ER) synthesize cytosolic and cytoskeletal proteins and proteins duced by these ribosomes are segregated during translation into
for import into the nucleus, mitochondria, and peroxisomes. the interior of the ER’s membrane cisternae.
Proteins that are to be incorporated into membranes, stored In both pathways misfolded proteins are conjugated to ubiqui-
in lysosomes, or eventually secreted from the cell are made on tin and targeted for proteasomal degradation.
Cytoplasmic Organelles 29
C H A P T E R
Nucleus
2
The Cytoplasm╇ ■╇ Cytoplasmic Organelles
Cisternae
Ribosomes
a c
Ribosomes Rough ER
Smooth ER
(a) The endoplasmic reticulum is an anastomosing network of The interconnected membranous cisternae of RER are flattened,
intercommunicating channels or cisternae formed by a continu- while those of SER are frequently tubular. (14,000X)
ous membrane, with some regions that bear polysomes appearing (c) In a very thin cultured endothelial cell, both ER (green) and
rough and other regions appearing smooth. While RER is the site mitochondria (orange) can be visualized with vital fluorescent
for synthesis of most membrane-bound proteins, three diverse dyes that are sequestered specifically into those organelles. This
activities are associated with smooth ER: (1) lipid biosynthesis, staining method with intact cells clearly reveals the continuous,
(2) detoxification of potentially harmful compounds, and lacelike ER present in all regions of the cytoplasm.
(3) sequestration of Ca++ ions. Specific cell types with well-developed (Figure 2–10c, © 2015 Thermo Fisher Scientific, Inc. Used under
SER are usually specialized for one of these functions. permission.)
(b) By TEM cisternae of RER appear separated, but they actually
form a continuous channel or compartment in the cytoplasm.
mRNA tRNA
5′ Signal sequence
SRP binds to SRP is is removed from
SRP receptor liberated 3′
growing polypeptide
New polypeptide
with initial
signal peptide SRP bound to
signal peptide
The newly translated amino terminus of a protein to be incorpo- ribosomal subunit, more firmly attaching the ribosome to the ER.
rated into membranes or sequestered into vesicles contains 15-40 The hydrophobic signal peptide is translocated through a protein
amino acids that include a specific sequence of hydrophobic pore (translocon) in the ER membrane, and the SRP is freed for
residues comprising the signal sequence or signal peptide. This reuse. The signal peptide is removed from the growing protein by
sequence is bound by a signal-recognition particle (SRP), which a peptidase and translocation of the growing polypeptide contin-
then recognizes and binds a receptor on the ER. Another recep- ues until it is completely segregated into the ER cisterna.
tor in the ER membrane binds a structural protein of the large
signal sequence is bound by a protein complex called the differences in the destinations of their major protein products
signal-recognition particle (SRP), which inhibits further and how these differences determine a cell’s histologic features.
polypeptide elongation. The SRP-ribosome-nascent peptide
complex binds to SRP receptors on the ER membrane. SRP
then releases the signal sequence, allowing translation to › ╺╺ MEDICAL APPLICATION
continue with the nascent polypeptide chain transferred to a Quality control during protein production in the RER and
translocator complex (also called a translocon) through the properly functioning ERAD to dispose of defective proteins
ER membrane (Figure 2–11). Inside the lumen of the RER, are extremely important and several inherited diseases result
the signal sequence is removed by an enzyme, signal pepti- from malfunctions in this system. For example, in some forms
dase. With the ribosome docked at the ER surface, translation of osteogenesis imperfecta bone cells synthesize and secrete
continues with the growing polypeptide pushing itself while defective procollagen molecules which cannot assemble
chaperones and other proteins serve to “pull” the nascent poly- properly and produce very weak bone tissue.
peptide through the translocator complex. Upon release from
the ribosome, posttranslational modifications and proper Smooth Endoplasmic Reticulum
folding of the polypeptide continue.
Regions of ER that lack bound polyribosomes make up the
RER has a highly regulated system to prevent nonfunc-
tional proteins being forwarded to the pathway for secretion smooth endoplasmic reticulum (SER), which is continuous
or to other organelles. New proteins that cannot be folded with RER but frequently less abundant (Figure 2–10). Lack-
or assembled properly by chaperones undergo ER-associated ing polyribosomes, SER is not basophilic and is best seen with
degradation (ERAD), in which unsalvageable proteins are the TEM. Unlike the cisternae of RER, SER cisternae are more
translocated back into the cytosol, conjugated to ubiquitin, tubular or saclike, with interconnected channels of various
and then degraded by proteasomes. shapes and sizes rather than stacks of flattened cisternae.
As mentioned, proteins synthesized in the RER can have SER has three main functions, which vary in importance
several destinations: intracellular storage (eg, in lysosomes in different cell types.
and specific granules of leukocytes), provisional storage in ■⌀ Enzymes in the SER perform synthesis of phospholipids
cytoplasmic vesicles prior to exocytosis (eg, in the pancreas, and steroids, major constituents of cellular membranes.
some endocrine cells), and as integral membrane proteins. These lipids are then transferred from the SER to
Diagrams in Figure 2–12 show a few cell types with distinct other membranes by lateral diffusion into adjacent
Cytoplasmic Organelles 31
2 C H A P T E R
The Cytoplasm╇ ■╇ Cytoplasmic Organelles
(a) Erythroblast (b) Eosinophilic leukocyte (c) Plasma cell (d) Pancreatic acinar cell
The ultrastructure and general histologic appearance of a cell are (c) Cells with extensive RER and a well-developed Golgi apparatus
determined by the nature of the most prominent proteins the cell show few secretory granules because the proteins undergo exocytosis
is making. immediately after Golgi processing is complete. Many cells, especially
(a) Cells that make few or no proteins for secretion have very little those of epithelia, are polarized, meaning that the distribution of RER
RER, with essentially all polyribosomes free in the cytoplasm. and secretory vesicles is different in various regions or poles of the cell.
(b) Cells that synthesize, segregate, and store various proteins in (d) Epithelial cells specialized for secretion have distinct polarity,
specific secretory granules or vesicles always have RER, a Golgi with RER abundant at their basal ends and mature secretory gran-
apparatus, and a supply of granules containing the proteins ready ules at the apical poles undergoing exocytosis into an enclosed
to be secreted. extracellular compartment, the lumen of a gland.
TV Vacuole
shipping
region
CF Secretory
vesicles
SV
TF
SV
Transport
Transport
vesicle
vesicle
Lumen of cisterna
filled with secretory
TV product
ER
G
M
b c
The Golgi apparatus is a highly plastic, morphologically complex (b) Morphological aspects of the Golgi apparatus are revealed
system of membrane vesicles and cisternae in which proteins and more clearly by SEM, which shows a three-dimensional snapshot
other molecules made in the RER undergo further modification of the region between RER and the Golgi membrane compart-
and sorting into specific vesicles destined for different roles in the ments. Cells may have multiple Golgi apparatuses, each with the
cell. general organization shown here and typically situated near the
(a) TEM of the Golgi apparatus provided early evidence about cell nucleus. (X30,000)
how this organelle functions. To the left is a cisterna of RER and (c) The Golgi apparatus location can be clearly seen in intact cul-
close to it are many small vesicles at the cis face (CF), or receiv- tured cells processed by immunocytochemistry using an antibody
ing face, of the Golgi apparatus, merging with the first of several against golgin-97 to show the many complexes of Golgi vesicles
flattened Golgi cisternae. In the center are the characteristic flat- (green), all near the nucleus, against a background of microfila-
tened, curved, and stacked medial cisternae of the complex. Cyto- ments organized as stress fibers and stained with fluorescent
logical and molecular data suggest that other transport vesicles phalloidin (violet). Because of the abundance of lipids in its many
(TV) move proteins serially through the cisternae until at the trans membranes, the Golgi apparatus is difficult to visualize in typical
face (TF), or shipping region, larger condensing secretory vesicles paraffin-embedded, H&E-stained sections. In developing white
(SV) and other vacuoles emerge to carry the modified proteins. blood cells with active Golgi complexes, the organelle can some-
elsewhere in the cell. Formation and fusion of the vesicles through times be seen as a faint unstained juxtanuclear region (sometimes
the Golgi apparatus is controlled by specific membrane proteins. called a “Golgi ghost”) surrounded by basophilic cytoplasm.
(X30,000) Inset: A small region of a Golgi apparatus in a 1-μm (Figure 2–13b reproduced, with permission from Naguro T, Iino A.
section from a silver-stained cell, demonstrating abundant glyco- Prog Clin Biol Res. 1989;295:250; Figure 2–13c, © 2015 Thermo Fisher
proteins within cisternae. Scientific, Inc. Used under permission.)
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prepared for this important step, he again refused. The principle manifested
in this refusal, is all the more worthy and memorable, because the poet had,
at this time, no hearthstone, no place that he could call his. His time was
employed, therefore, in travelling about, from place to place, and from friend
to friend; now to good Robert Jones, in Wales,—the man whom De Quincy
conjectures to have had no brains, and for which I owe the said De Quincy a
grudge, notwithstanding that I think more highly of him than any other man
now living in these realms,—and now to Mr. Rawson’s, of Millhouse,
Halifax, who had married Wordsworth’s cousin, Miss Threlkeld, the lady
who brought up Dorothy Wordsworth, the dearly beloved sister of the poet.
In 1794, Wordsworth writes to his friend Mathews “that his sister is under
the same roof with him; but that he is doing nothing, and knows not what
will become of him.” All his path lay dark and gloomy before him. He was
recommended to study the law, but he absolutely refused; and the Fates
seemed to be sporting with him. His love for Dorothy grew in him every day,
and it was in the year I am now speaking of, that she, having accompanied
him by coach from Halifax to Kendal, walked with him from the latter place
to Grasmere—eighteen miles, and from thence to Keswick—fifteen miles
further, where they put up at a farm-house, called Windybrow, and became
thenceforth all-in-all to each other.
But the grand question for Wordsworth now to solve was, how he should
earn his daily bread. His poetry brought no grist to the mill; and he had no
friends to fall back on. In this condition, he wrote to his friend Mathews,
who was connected with the London Press, to get him employment upon one
of the daily or weekly papers, but without success. He projected, likewise,
several literary journals, with the same bad fortune.
At last, however, Providence had ordained that a young man, named
Raisley Calvert, should die, and leave the poet £900. Poor fellow! He seems
to have been born for this special purpose. Wordsworth had known him
previous to his illness, and the young man was so impressed with the genius
and capabilities of the poet, that he bequeathed his little fortune to him, in
order that he might have leisure to develope them. This was in 1795, whilst
the negociations respecting the newspaper employment were still pending. It
was at Mrs. Sowerby’s hostel, at the sign of the “Robin Hood,” in Penrith,
where poor Calvert lay sick—where Wordsworth nursed and tended him,—
and where also he died.
This bequest was of the last importance to Wordsworth, for it rescued him
from poverty and distress, and enabled him to live.
All the avenues of the world were closed against him; for he was, by
nature and education, unfitted for the tradesman’s service, or the clerk’s
office, or the schoolman’s desk. He was a solitary dreamer; a lonely,
meditative man, who thought golden thoughts, and built starry ladders to
heaven, and did all sorts of strange things, similar in character, which
unfortunately could not be sold as wares in the public market-place. And it
was necessary, if Wordsworth was to be any thing, that he should continue
this foolish habit of dreaming, of rhyming, and writing, into which he had so
singularly fallen. For in her education of the poet, Nature—who is a wise
teacher and developer—always adopts one regular method, although she
frequently varies the process: and this method consists in impressing the
mind with her manifold forms, colours, sounds, and works, that they may
hereafter be reproduced in the glorious imagination of the poet, and shine,
like rich mosaics, in the wisdom of his teaching. But for the friend alluded
to, however, our poet could never have afforded to have gone through his
initiation in the mysteries of poetry, but must have squatted down as a
school-master in some suitable town (which De Quincy says he once thought
of doing), and have turned professor of flogging Greek into pigmy
humanities. Or, perhaps, the Poet of Rydal might have been little better than
a scribe-engro, if I may coin a word from George Borrows’ “Romany,” and
have ended his life as a political or literary hack, in some Times’ omnibus, or
other vehicle of less note and more principle.
Fortunately for literature, and for men, as the readers of literature, Raisley
Calvert was born and died—and the “Excursion” was written. I would not,
however, be thought to speak ungenerously of poor Calvert:—God forbid!—
but still I cannot help thinking about Providence, and His dark, inscrutable
ways how He smites one frail child to the grave that another may have
leisure to sing songs.—The poet never forgets the bounty and generosity of
the poor dead heart, now hushed to silence, but raises this monument to his
memory:
“Calvert! it must not be unheard by them
Who may respect my name, that I to thee
Owe many years of early liberty.
This care was thine, when sickness did condemn
Thy youth to hopeless wasting, root and stem,
That I, if frugal and severe, might stray
Where’er I liked, and finally array
My temples with the Muse’s diadem.
Hence, if in freedom I have loved the truth,
If there be aught of pure, or good, or great,
In my past verse, or shall be in the lays
Of highest mood, which now I meditate,
It gladdens me, O worthy short-lived youth,
To think how much of this will be thy praise.”
maintained for him a saving intercourse with his true self. It was to her that
he was indebted for many salutary admonitions; it was she who, in the
midst of the clashing politics and noisy-throated revolutions of Europe,
preserved him “still a poet,” and made “him seek beneath that name, and
that alone, his office upon earth.”
In 1795, Wordsworth, accompanied by his sister, left Cumberland, for
Racedown Lodge, near Cremkerne, in Dorsetshire. The country was
delightful, and the house pleasantly situated, with a garden attached to it.
They passed their time in reading, gardening, writing, and in translating
Ariosto, and other Italian poets. We also find the poet making imitations of
Juvenal’s Satires, copies of which he sent to his friend Wrangham, with a
view to joint publication. They were never printed, however; and
Wordsworth, in 1805, when he was urged by the same friend to allow them
to appear, repudiated them altogether, and regretted that he had spent so
much time in their composition, declaring that he had come to a “fixed
resolution to steer clear,” now and for ever, of all personal satire. During
this same year of ’95, he finished his poem on “Guilt and Sorrow,” and
began his tragedy of “The Borderers,” which was completed before the
close of the following year. It is a cumbrous affair, and will not act; but it
contains some fine passages. It was first published in 1842, and was offered
to Mr. Harris, the manager of the Covent Garden Theatre, through Mr.
Knight, the actor, and was, as Wordsworth confesses, “judiciously returned,
as not calculated for the stage.”
In the year 1797, Coleridge came to Racedown. Miss Wordsworth
describes him in one of her letters as “a wonderful man;” his conversation
teeming with “soul, mind, and spirit,”—three things very nearly related to
each other, one would think. He is benevolent, too, good-tempered, and
cheerful, like her dear brother William, and “interests himself much about
every little trifle.” At first she thought him very plain—that is for about
“three minutes” for he is “pale, thin, has a wide mouth, thick lips, and not
very good teeth; longish, loose growing, half-curling, rough black hair.” But
he no sooner began to talk than she forgot his want of comeliness, his bad
teeth, and wide mouth, and was entranced by the magic of his eloquence.
“His eye,” she adds, “is large and full, not very dark, but grey; such an eye
as would receive from a heavy soul the dullest expression; but it speaks
every emotion of his animated mind; it has more of the ‘poet’s eye in a fine
frenzy rolling,’ than I ever witnessed. He has fine dark eye-brows, and an
overhanging forehead.”
Such was the apparition of Coleridge, in Wordsworth’s house at
Racedown. The poet himself was delighted with his visitor, and they were
soon in deep conversation about literature, and their own several
adventures, and proposed argosies on that great and shoreless deep.—
Wordsworth read a new poem, which he had just written, called the “Ruined
Cottage,”—and Coleridge praised it highly. Then they sat down to tea, and
presently the latter “repeated two acts and a half of his tragedy, ‘Osoris.’ ”
The next morning, the “Borderers” was read, and passages from “twelve
hundred lines of blank verse,—superior,” says Coleridge, in a letter to a
friend, “I hesitate not to aver, to anything in our language which in any way
resembles it.” “I speak with heart-felt sincerity, and I think combined
judgment, when I tell you,” he elsewhere writes, “that I feel a little man by
his side.”
Coleridge came several times to see his big man, after this; and the two
poets grew so much attached to each other, and found such profitable
advantages in each other’s conversation and literary judgments, that they
resolved to dwell nearer to each other. Accordingly Wordsworth and his
sister went to live at Alfoxden, near Stowey, where Coleridge was residing.
This was in July, 1797,—and he describes his sojourn there as a very
“pleasant and productive time of his life.” The house which Wordsworth
occupied belonged to Mr. St. Aubyn, who was a minor,—and the condition
of occupancy seems to have been, that the poet should keep the house in
repair.
In one of Miss Wordsworth’s letters, dated August 14, 1797, she speaks
with great enthusiasm and delight, both of the house and the country. “Here
we are,” she says, “in a large mansion, in a large park, with seventy head of
deer around us.... Sea, woods wild as fancy ever painted, brooks clear and
pebbly as in Cumberland, villages so romantic, &c. The woods are as fine
as those at Lowther, and the country more romantic; it has the character of
the less grand part of the neighbourhood of the Lakes.” She then describes
their “favourite parlour,” which, like that of Racedown, looks into the
garden. They were three miles from Stowey, and only two from the sea.
Look which way they would, their eyes were filled with beauty: smooth
downs, and valleys with small brooks running down them, through green
meadows, hardly ever intersected with hedges, but scattered over with trees.
The hills were covered with bilberries, or oak woods. They could walk for
miles over the hill-tops, which was quite smooth, without rocks.
And in this beautiful locality did the poet reside for about twelve
months, composing during that time, all the poems contained in the first
edition of the “Lyrical Ballads,” with the exception of the “Female
Vagrant.” These Ballads are in many ways remarkable: in the first place,
because they were the joint production of men who subsequently proved
themselves to be two of our greatest poets; and, secondly, because they
brought these men prominently before the eyes of the public and of the
critical world. The Ballads, as they were called, were likewise of a very
high order; and it is not too much to say, that such a book of poems as this
had not been published since the Augustan era of our literature, Milton’s
alone excepted, if Milton may not be said to have closed that era. Here first
appeared the “Ancient Mariner,” and the “Nightingale,” by Coleridge;
“Tintern Abbey,” and “Lines left under a Yew-tree Seat,” by Wordsworth;
four poems which of themselves were sufficient to float half a dozen
volumes. It is true that the “Ancient Mariner,” the “Old Navigator,” as
Coleridge loved to call it, is what may be styled a made-up poem—a wild,
unearthly patchwork of the imagination,—but it contains, nevertheless, such
passages as it would be rare to match outside those seas. It is full, too, of all
kinds of music—sweet, wild, natural, and supernatural—now grand, like
the rolling bass of some mighty organ, and now, ærial, celestial; catching up
the reader into a strange heaven, and filling him with an unspeakable
ecstacy. Wonderful power is likewise manifested in the structure of the tale;
and one is amazed how so slender an incident, as that upon which the tale is
founded, could be worked out so successfully, and with such deep and
thrilling interest. The “Nightingale,” however, is quite a different poem, and
is redolent of nature. “Tintern Abbey,” and “Lines left under a Yew-tree
Seat,” are in Wordsworth’s best style, and have never been surpassed by
him, in the fullest maturity of his genius.
The idea of the “Ballads” originated in the following circumstances:
Wordsworth and his sister, accompanied by Coleridge, commenced a
pedestrian tour, in November 1797, to Linton, and the Valley of Stones,
near it. The whole party, however, were so poor that they could ill afford the
expense of the journey, and the two poets resolved to write a poem for the
“New Monthly Magazine,” for which they hoped to get £5, and thus
balance the outlay which they required for the tour. The course of the
friends lay along the Quartock Hills, towards Watchet; and here it was that
Coleridge planned his “Old Navigator,” the base of it being, as he said, a
dream of Cruikshanks’. Wordsworth and Coleridge were to have written
this poem conjointly, but the great dissimilarity of their manner soon
compelled them to abandon this idea, and Coleridge was left to complete
the work by himself. Wordsworth suggested, however, as some crime was
to be committed by the Mariner, which was to bring upon him a spectral
persecution in his wanderings, as the consequence of that crime, that he
should be represented as having killed an Albatross on entering the South-
Sea, and that the tutelary spirits of these regions should follow him and
avenge the crime. The navigation of the ship by the dead men was also a
suggestion of Wordsworth’s. As Coleridge proceeded with his work, it was
very soon found that it would be too long for the Magazine, and they began
to think of issuing it as a volume, along with other poems, by both bards.
These poems were to be founded “on supernatural subjects, taken from
common life, but to be looked at, as much as might be, through an
imaginative medium.” Wordsworth’s share in the poetical contributions to
this volume, besides those already mentioned were, amongst others, “The
Idiot Boy,” “Her Eyes are Wild,” “We are Seven,” and “The Thorn.” The
last verse of “We are Seven,” was composed first, and Coleridge threw off,
impromptu, the first verse of the poem, whilst the little party were sitting
down to tea, in the pretty little parlour at Alfoxden, which looked out into
the garden. Speaking of the “Idiot Boy,” Wordsworth says:—“The last
stanza, ‘The cocks did crow, and moon did shine so cold,’ was the
production of the whole. The words were reported to me by my dear friend
Thomas Poole; but I have since heard the same reported of other idiots. Let
me add, that this long poem was composed in the groves of Alfoxden,
almost extempore; not a word, I believe, being corrected, though one stanza
was omitted. I mention this in gratitude to those happy moments, for in
truth I never wrote anything with so much glee.”
It was in 1798 that the Lines to Tintern Abbey were written. The poet
and his sister had been staying for a week with Mr. Cottle, of Bristol,
visiting Coleridge by the way, who had a little time before resigned his
ministerial engagement with a Unitarian congregation at Bristol, and was
now in receipt of an annuity of £150, given to him by the magnificent
generosity of “Josiah and Thomas Wedgwood.” From Mr. Cottle’s they
proceeded to the Banks of the Wye, crossed the Severn ferry, and walked
ten miles further to Tintern Abbey, a very beautiful ruin on the Wye. They
proceeded, next morning, along the river, through Monmouth, to Goderich
Castle, returning to Tintern in a boat, and from thence in a small vessel back
again to Bristol.
“The Wye,” says Wordsworth, “is a stately and majestic river, from its
width and depth, but never slow and sluggish—you can always hear its
murmur. It travels through a woody country, now varied with cottages and
green meadows, and now with huge and fantastic rocks.... No poem of mine
was composed under circumstances more pleasant for me to remember than
this [viz., “Tintern Abbey:”]—I began it upon leaving Tintern, after
crossing the Wye, and concluded it just as I was entering Bristol, in the
evening, after a ramble of four or five days with my sister. Not a line of it
was uttered, and not any part of it written down, till I reached Bristol. It was
published almost immediately after in the “Lyrical Ballads.”
“Peter Bell” was likewise written about this time; and the following
interesting particulars, respecting its origin, are furnished by the poet:
“This tale was founded upon an anecdote which I read in a newspaper, of
an ass being found hanging his head over a canal, in a wretched posture.
Upon examination, a dead body was found in the water, and proved to be
the body of its master. In the woods of Alfoxden, I used to take great delight
in noticing the habits, tricks, and physiognomy of asses; and it was here, no
doubt, that I was put upon writing the Poem of ‘Peter Bell,’ out of liking for
the creature that is so often dreadfully abused. The countenance, gait, and
figure of Peter were taken from a wild rover with whom I walked from
Builth, on the river Wye, downwards, nearly as far as the town of Hay. He
told me strange stories. It has always been a pleasure to me through life, to
catch at every opportunity that has occurred in my rambles, of being
acquainted with this class of people. The number of Peter’s wives was taken
from the trespasses in this way of a lawless creature who lived in the county
of Durham, and used to be attended by many women, sometimes not less
than half a dozen, as disorderly as himself; and a story went in the county,
that he had been heard to say, whilst they were quarrelling: “Why can’t you
be quiet?—there’s none so many of you.’ Benoni, or the child of sorrow, I
knew when I was a school-boy. His mother had been deserted by a
gentleman in the neighbourhood, she herself being a gentlewoman by birth.
The circumstances of her story were told me by my dear old dame, Ann
Tyson, who was her confidante. The lady died broken-hearted. The crescent
moon, which makes such a figure in the prologue, assumed this character
one evening, while I was watching its beauty, in front of Alfoxden House. I
intended this poem for the volume before spoken of; but it was not
published for more than twenty years afterwards. The worship of the
Methodists, or Ranters, is often heard during the stillness of the summer
evening, in the country, with affecting accompaniments of moral beauty. In
both the psalmody and voice of the preacher there is not unfrequently much
solemnity, likely to impress the feelings of the rudest characters, under
favourable circumstances.”
It was during Wordsworth’s residence in the South, in 1794, that a
circumstance occurred, which has not been alluded to before in these
memoirs, but which is interesting in itself, and still more so from the fact
that it was the means of making Wordsworth acquainted with Coleridge,
Southey, Robert Lovell, and George Burnet. I will relate this circumstance
in the language of a writer for Chambers’ Papers for the People, and quote
still further passages from the same tract, illustrative of the life of
Wordsworth and his friends, at this time.[G]
The circumstance was as follows: Coleridge, Southey, Lovell, and
Burnet “came down to Bristol, as the most convenient part from which they
could embark for the wild banks of the Susquehana. On that remote river
they were to found a Platonic Republic, where everything was to be in
common, and from which vice and selfishness were to be for ever excluded.
These ardent and intellectual adventurers had made elaborate calculations
how long it would take them to procure the necessaries of life, and to build
their barns, and how they should spend their leisure in what Coleridge sung
as
Yet, it is supposed, they knew nothing of the Susquehana more than of any
other American river, except that its name was musical and sonorous; and
far from having anything wherewith to convey themselves and their
moveables across the Atlantic, they had to borrow five pounds to make up
their lodging bill. This sum was advanced them, with unalloyed pleasure,
by Mr. Cottle, a bookseller in the town, a benevolent and worthy man, who
seems almost to have been located there for no other purpose than to
introduce the three chief Lake Poets to the world.
“The bubble of the Susquehana, or, as it was called, Pantisocracy, was
exploded, by Southey, Coleridge, and Lovell, all getting into the bonds of
matrimony, which have a miraculous virtue in testing the solidity of
schemes of life. They married three sisters of the name of Fricker. It was the
perpetual restlessness of Coleridge which first brought him and his
companions into contact with Wordsworth. The former wonderful man, in
capabilities perhaps the mightiest of that illustrious group, and in his mental
constitution one of the most puzzling psychological phenomena which
human nature has ever presented, was the originator of the Pantisocratic
proposal. He was a man of luxurious imagination, deep emotiveness,
various learning, and an exquisite nervous susceptibility. In 1795, he was
making excursions through the lovely and tranquil scenery of
Somersetshire, when he became acquainted with a most worthy and
excellent man, Mr. Poole, resident in the quiet village of Stowey. On his
return to Bristol, where he was married, he still exhibited his uneasiness.
First he removed to his immortal rose-bound cottage at Clevedon, then back
to the pent-up houses at Redcliff Hill, and from these again to the more
open situation of Kingsdown. Nothing would then satisfy him but he must
set up a serial, to be called ‘The Watchman;’ and his own sketches of his
travelling canvass for that periodical, might take rank with some chapters of
Don Quixote. Take, for instance, this picture of a great patriot at
Birmingham, to whom he applies for his magnificent patronage:—He was
‘a rigid Calvinist, a tallow-chandler by trade. He was a tall, dingy man, in
whom length was so predominant over breadth, that he might almost have
been borrowed for a foundry poker! Oh, that face!—I have it before me at
this moment. The lank, black, twine-like hair, pinguinitescent, cut in a
straight line along the black stubble of his thin, gunpowder eyebrows, that
looked like a scorched aftermath from a last week’s shaving. His coat-collar
behind, in perfect unison, both of colour and lustre, with the coarse yet glib
cordage which I suppose he called his hair, and which, with a bend inward
at the nape of the neck—the only approach to flexure in his whole figure—
slunk in behind his waistcoat; while the countenance, lank, dark, very hard,
and with strong, perpendicular furrows—gave me a dim notion of some one
looking at me through a gridiron,—all soot, grease, and iron.’ This
thoroughbred lover of liberty, who had proved that Mr. Pitt was one of the
horns of the second beast in the ‘Revelations’ that spake as a dragon,
nevertheless declined to take ‘The Watchman’; and in short, after a
disastrous career, that serial died a natural death. The disappointed editor
took refuge, for a brief season, with Mr. Poole, at Stowey, and there, for the
first time, he met Wordsworth, who then resided about twenty miles off, at
Racedown, in Dorsetshire.
“Coleridge returned for a short time to Bristol, but in January, 1797, he
removed to Stowey, where he rented a small cottage. This must have been a
pleasant episode in the lives of the gifted individuals whom it brought
together in that sweet village. Wordsworth, who was now twenty-seven, had
come with his sister to Alfoxden, which was within two miles of Stowey.
Charles Lloyd, a young man of most sensitive and graceful mind, and of
great poetical susceptibility, resided in the family with Coleridge. Charles
Lamb, then in the spring-time of his life, was also a frequent inmate; and
often afterwards, under the cloud which lowered over his noble devotedness
in London, his fancy wandered back to that happy valley.—Why, says he to
Charles Lloyd, who unexpectedly looked in upon him in the great Babylon:
The Pantisocratist, George Burnet, was also a visitor. Mrs. Coleridge herself
had a poetical taste, and there is one very graceful piece of hers written on
the receipt of a thimble from her kind friend Mr. Cottle. Just such a thimble,
sings Sarah Coleridge—
Hartley Coleridge, the ærial child who awakened the fears and sympathies
of Wordsworth, was a fine boy, rejoicing his parents’ hearts; and the happy
pair had cut a road into their neighbours’ orchards, that they might pass to
their firesides under the arches of blossoms, with a speed suiting to their
affections. Alas! that sweet Stowey. Cottle, in his old age, has painted one
or two pictures of it and of its gifted habitants, now in their graves, that go
to the heart. Take the scene with Coleridge in the jasmine arbour, where the
tripod table was laden with delicious bread and cheese, and a mug of the
true brown Taunton ale. ‘While the dappled sunbeams,’ says the old man,
calling up kindly memories, ‘played on our table through the umbrageous
canopy, the very birds seemed to participate in our felicities, and poured
forth selectest anthems. As we sat in our sylvan hall of splendour, a
company of the happiest mortals, the bright blue heavens, the sportive
insects, the balmy zephyrs, the feathered choristers, the sympathy of
friends, all augmented the pleasurable to the highest point this side the
celestial.... While thus elevated in the universal current of our feelings, Mrs.
Coleridge approached with her fine Hartley; we all smiled, but the father’s
eye beamed transcendental joy. But all things have an end! Yet pleasant it is
for Memory to treasure up in her choicest depository a few such scenes
(those sunny spots in existence), on which the spirit may repose when the
rough adverse winds shake and disfigure all besides.’ Or take the more
lively visit to Alfoxden, on Wordsworth’s invitation. Away they all went
from Stowey; the poet and Emmeline, Coleridge and Cottle. They were to
dine on philosopher’s fare—a bottle of brandy, a loaf, a piece of cheese, and
fresh lettuces from Wordsworth’s garden. The first mishap was a theftuous
abstraction of the cheese; and, on the back of it, Coleridge, in the very act
of praising the brandy as a substitute, upset the bottle, and knocked it to
pieces. They all tried to take off the harness from the horse. Cottle tried it,
then the bard of Rydal; but in vain. Coleridge, who had served his
apprenticeship as Silas Comberbatch in the cavalry, then twisted the poor
animal’s neck almost to strangulation; but was at last compelled to
pronounce that the horse’s head must have grown since the collar was put
on! It was useless, he said, to try to force so huge an os frontis through so
narrow a collar. All had given up, when lo! the servant girl turned the collar
upside down, and slipped it off in an instant, to the inconceivable wonder
and humiliation of the poets, who proceeded to solace themselves with the
brown bread, the lettuces, and a jug of sparkling water. Who, knowing the
subsequent fate of the tenants of Stowey, would not love to dwell on these
delightful pictures of their better days?
“It must not be supposed, however, that the tempter never entered into
this Eden; but when he did so, it was generally through the mischief-making
pranks of Coleridge, who constantly kept his friends in hot water. He and
Lamb had just published a joint volume of poems, and Coleridge could not
refrain from satirising and parodying their offspring in the newspapers.
Take this epigram as a specimen:—
——“a tune
Harsh and of dissonant mood from their complaint.”
One of the party was a Dane, a vain and disgusting coxcomb, whose
conversation with Coleridge, whom he first took for a ‘Doctor Teology,’
and then for ‘un philosophe,’ actually outburlesqued burlesque. The
astounded bard, for the first time in his life, took notes of a dialogue, of
which a single sample is enough.
“The Dane.—Vat imagination! vat language! vat vast science! vat eyes!
vat a milk white forehead! Oh my heafen! vy, you’re a got!
“Answer.—You do me too much honour, sir.
“The Dane.—Oh me, if you should tink I is flattering you! I haf ten
tousand a year—yes, ten tousand a year—yes, ten tousand pound a year!
Vell, and vat is dhat? Vy, a mere trifle! I ’ould’nt gif my sincere heart for ten
times dhe money! Yes, you’re a got! I a mere man! But, my dear friend,
dhink of me as a man! Is—is—I mean to ask you now, my dear friend—is I
not very eloquent? Is I not speak English very fine?
“And so his Daneship, in this extraordinary style, went on fishing for
compliments, and asking whether he did not speak just like Plato, and Cato,
and Socrates, till he lost all opinion of Coleridge on finding that he was a
Christian. The discarded poet then wrapped himself in his great-coat, and
looked at the water, covered with foam and stars of flame, while every now
and then detachments of it ‘darted off from the vessel’s side, each with its
own constellation, over the sea, and scoured out of sight like a Tartar troop
over a wilderness.’ By and by he lay down, and ‘looking up at two or three
stars, which oscillated with the motion of the sails, fell asleep.’
“They landed at Hamburg, on the Elbe Stairs, at the Boom-House.
Wordsworth, with a French emigrant, whose acquaintance he had cultivated
at sea, went in search of a hotel, and put up at ‘Die Wilde Man,’ while the
other wild man, Samuel Taylor Coleridge, strolled about, amusing himself
with looking at the ‘Dutch women, with large umbrella hats shooting out
half a yard before them, and a prodigal plumpness of petticoat behind,’ and
many similar striking and unusual spectacles.
“In Hamburg the pair were introduced to the brother of the poet
Klopstock, and to Professor Ebeling, a lively and intelligent man, but so
deaf that they had to ‘drop all their pearls into a huge ear-trumpet.’ At Mr.
Klopstock’s they saw a bust of the poet, whom they afterwards visited. It
had a solemn and heavy greatness in the countenance, which corresponded
with the notions entertained by Coleridge of his style and genius, and which
were afterwards discovered not to exist in the prototype himself. Coleridge,
whose chief object in coming to Germany was to become acquainted with
the German language and literature, left Wordsworth in Hamburg, and went
to Ratzeburg, where he boarded in the pastor’s house. He returned,
however, for a few days, to take final leave of his friend, and the two paid a
visit to Klopstock together. His house was one of a row of what appeared
small summer-houses, with four or five rows of young meagre elms in
front, and beyond these a green, bounded by a dead flat. The bard’s
physiognomy disappointed them as much as his domicile. Coleridge
recognised in it no likeness to the bust, and no traces either of sublimity or
enthusiasm. Klopstock could only speak French and German, and Coleridge
only English and Latin, so that Wordsworth, who was accomplished in
French, acted as interpreter. It may here be mentioned that this ignorance of
Coleridge’s brought upon him a peculiar sort of civility at Ratzeburg. The
amtmann of that place, anxious to be civil, and totally unable to find any
medium of communication, every day they met, as the only courtesy he had
it in his power to offer, addressed to him the whole stock of English he
possessed, which was to this effect:— ‘——ddam your ploot unt eyes, my
dearest Englander, vhee goes it?’ The conversation with Klopstock turned
entirely upon English and German literature, and in the course of it
Wordsworth gave ample proofs of his great taste, industry, and information,
and even showed that he was better acquainted with the highest German
writers than the author of the ‘Messiah’ himself. On his informing the latter
that Coleridge intended to translate some of his odes, the old man said to
Coleridge—‘I wish you would render into English some select passages of
the “Messiah,” and revenge me of your countrymen.’ ‘This,’ says
Coleridge, ‘was the liveliest thing he produced in the whole conversation.’
That genius was, however, deeply moved, but could not help being
disgusted with the venerable bard’s snow-white periwig, which felt to his
eye what Mr. Virgil would have been to his ear. After this, Coleridge left
Hamburg, and resided four months in Ratzeburg, and five in Gottingen.
Wordsworth had two subsequent interviews with Klopstock, and dined with
him. He kept notes of these conversations, some of which are given in
‘Satyrane’s Letters,’ in the second volume of the ‘Biographia Literaria.’
One or two incidents strongly illustrate Wordsworth’s peculiar character
and poetical taste. He complained, for example, of Lessing making the
interest of the ‘Oberon’ turn upon mere appetite. ‘Well, but,’ said
Klopstock, ‘you see that such poems please everybody.’ He immediately
replied, that ‘it was the province of a great poet to raise a people up to his
own level—not to descend to theirs.’ Klopstock afterwards found fault with
the Fool in ‘Lear,’ when Wordsworth observed that ‘he gave a terrible
wildness to the distress’—a remark which evinced a deep appreciation of
that awful drama. Wordsworth subsequently made a short tour, and visited
Coleridge at Gottingen on his return.’
Wordsworth, during the greater part of his time in Germany lived at
Goslar, and he found the people neither very friendly nor hospitable. Goslar
is situate at the foot of the Hartz Mountains, which are covered with fine
oaks and beech, and the poet and his sister, to make up for the loss of
society in the town, sought solitude amongst these magnificent woods.
Among the poems written at this time were, “Strange fits of passion have I
known,” “Three years she grew in sun and shower,” “Lines to a Sexton,”
“The Danish Boy,” intended as a prelude to a ballad never written; “A
Poet’s Epitaph,” “Art thou a Statist?” “Lucy Gray.” All these poems and
many more were written in 1799; and the latter is founded on a
circumstance related by the poet’s sister “of a little girl, who, not far from
Halifax, in Yorkshire, was bewildered in a snow-storm. Her footsteps were
tracked by her parents to the middle of the lock of a canal, and no other
footsteps of her, backward or forward, could be traced. The body, however,
was found in the canal. The way in which the incident was treated, and the
spiritualizing of the character, might furnish hints for contrasting the
imaginative influences, which I have endeavoured to throw over common
life, with Crabbe’s matter-of-fact style of handling subjects of the same
kind. This is not spoken to his disparagement,—far from it; but to direct the
attention of thoughtful readers into whose hands these notes may fall, to a
comparison that may enlarge the circle of their sympathies, and tend to
produce in them a catholic judgment.” “Lines written in Germany, ’98-9”
have the following note attached to them in the “Memoirs,” with which I
will conclude these extracts.
“A bitter winter it was when these verses were composed, by the side of
my sister, in our lodgings at a draper’s house, in the romantic imperial town
of Goslar, on the edge of the Hartz Forest. In this town the German
emperors of the Franconian line were accustomed to keep their court, and it
retains vestiges of ancient splendour. So severe was the cold of this winter
that when we passed out of the parlour warmed by the stove, our cheeks
were struck by the air as if by cold iron. I slept in a room over a passage
that was not ceiled. The people of the house used to say, rather unfeelingly,
that they expected I should be frozen to death some night; but with the
protection of a pelisse lined with fur, and a dog-skin bonnet, such as was
worn by the peasants, I walked daily on the ramparts, in a sort of public
ground or garden, in which was a pond. Here I had no companion but a
kingfisher, a beautiful creature that used to glance by me. I consequently
became much attached to it.” During these walks he composed the ‘Poet’s
Epitaph.’ His ‘Poem of Ruth,’ ‘The Address to the Scholars of a Village
School,’ ‘The two April Mornings,’ ‘The Fountain,’ ‘A Conversation,’
‘Matthew,’ and a variety of others were likewise written about this time. In
his correspondence with Coleridge at this time, the latter speaks in terms of
the highest affection for him. ‘I am sure I need but say,’ he writes, ‘how you
are incorporated with the better part of my being; how whenever I spring
forward into the future with noble affections, I always alight by your
side.’ ”
On the 10th of February, 1799, Wordsworth and his sister left Goslar,
and returned towards England. The poet was now nearly thirty years of age;
and as the gates of the Imperial City closed behind him, he felt like a bird
suddenly released from captivity, and resolved to build up some stately
architecture of verse, which men would not willingly let die. Accordingly
he commenced the “Prelude,” within the very hum of the city. Six out of the
fourteen books which compose it, were written before 1805.
In the spring of 1799 the poet and his sister returned to England; and in a
letter to Cottle, written immediately after their arrival, we find them in “the
county of Durham, just on the borders of Yorkshire,” thankful, after
sufficient experience of Germany, for the dear face of old England once
more.