THE PHOTORECEPTORS OF THE NURSE SHARK,

GINGLYMOSTOMA CIRRATUM AND THE

STING RAY, DASYATIS SAyr
D. 1. HAMASAKI
Bascom Palmer Eye Institute, University of Miami
AND
S. H. GRUBER2
Institute of Marine Science, University of Miami

ABSTRACT
A light microscopic study of the retina of the nurse shark, Gingly-
mostoma cirratum, and the sting ray, Dasyatis sayi, demonstrated two
types of receptors which had the main morphological characteristics of
rods and cones. The rods were very similar in both species but the cones
differed slightly-those of Dasyatis were more cone-like than those of
Ginglymostoma. The ratio of rods to cones was 7: 1 for Ginglymostoma
and 5: I for Dasyatis.

INTRODUCTION

In a recent study of the retina of the lemon shark, Negaprion breviros-

tris, two types of photo receptors were recognized by their differences in
shape and size (Gruber, Hamasaki, & Bridges, 1963). The ones iden-
tified morphologically as rods were tall and cylindrical with large outer
segments, while the others, identified as cones, Were short and conical
with small tapered outer segments (see Table 1 for measurements). In
the posterior pole of the eye, the rods were more numerous than the cones
by a ratio of 12: 1.
Cones have been described in other elasmobranchs (Mustelus, Mylio-
batis, Squatina, and Lamna), and one notable feature has been their
variability in shapes (see Walls, 1942; Rochon-Duvigneaud, 1943); those
of Myliobatis were completely cone-like while those of Mustelus were
more rod-like. This variation in shapes has led Walls to postulate that the
cones of elasmobranchs are "new," derived secondarily from the rods.
In a concurrent study of the electrical responses from the retina--elec-
troretinogram (ERG )-of three elasmobranchs, Negaprion brevirostris
(lemon shark), Ginglymostoma cirratum (nurse shark), and Dasyatis
sayi (sting ray), it was found that the shape and properties of the ERG's
were similar in all three species (Hamasaki & Bridges, 1964). There
was no evidence of a "break" in the dark-adaptation curves in the three
species which might indicate separate photopic and scotopic mechanisms
determining retinal sensitivities. However, the shapes of the ERG's were
)This inveS'ligationwas supported in part by a Public Health Service Research Grant,
NB 04630-2 from the National Institutes of Neurological Diseases and Blindness, and
Officeof Naval Research Contract Task Order Nom 4008(10).
2ContributionNo. 660 from The Institute of Marine Science, University of Miami.
1052 Bulletin of Marine Science [15(4)
markedly different depending on whether the animals were dark- or
light-adapted.
Thus it seemed of interest to compare the retinas of Ginglymostoma
and Dasyatis with that of Negaprion to see if they reflected the similarities
found in the electrical responses from the eye. In addition it was hoped
that some evidence might be gained on the question of the evolution of
the cones in the elasmobranchs. It will be shown that two types of photo-
receptors were present in both Ginglymostoma and Dasyatis which were
easily differentiated morphologically into rods and cones at the light
microscope level. The cones differed slightly from each other and from
those of the N egaprion.
We thank Mr. O. Navarro for his excellent preparation of the histological
materials, Dr. W. J. Wisby for his encouragement, and the Florida Lions'
Eye Bank for the use of its facilities.

FIGURE1. Vertical section of the retina of Ginglymostoma cirratum. Four cones

can be seen in this section: oil immersion, Mallory's Triple Stain. p.e.-pigment
epithelium; I.r.c.-Iayer of rods and cones; e.l.m.-external limiting membrane;
o.n.I.-outer nuclear layer; e-ellipsoid; p-parabaloid.
 1965] Hamasaki & Gruber: Eiasmobranch Photoreceptors 1053
METHODS

The animals were taken on a bright sunny day in the Florida Keys
by harpooning. They were killed and the enucleated eyes placed im-
mediately in Kolmer's fixative. Best fixation was obtained by making
a cut in the cornea to allow the fixative to penetrate into the eye.
Routine paraffin sections of 4 to 5 JL were made, and the retinas were
stained with various stains, but the best differentiation of the receptors was
obtained with Mallory's Triple Stain. Both vertical and tangential sections
were studied.
OBSERVATIONS

Ginglymostoma cirratum.-Eyes from three young adult specimens (total

length 90, 117, and 132 em) were examined. In all cases, two types
of photo receptors were recognized by their difference in shape, size,
and staining properties. A photomicrograph of the photoreceptors from
the posterior pole of the eye of a Gingiymostoma is shown in Figure 1,
and their sizes are recorded in Table I.
The rods were more or less constant in size and shape throughout the
retina and had an overall length of 33 JL. The outer segments were large
and cylindrical and extended into the pigment epithelial cells. The inner
segments of the rods were 16 JL in length and consisted of the heavily
stained ellipsoid and the lighter but evenly stained parabaloid. The aver-
age diameter of the outer segment was 2 JL while that of the inner seg-
ment was 2.5 JL

TABLE 1
A VERAGEMEASUREMENTS AND COUNTS OF RODS AND
CONES FROM POSTERIOR POLE OF THE EVE
---
Rods Cones
Outer Inner Outer Inner Rod:

-S
segment
..
<1l
.•...•
segment
..c::
..
<1l
.•...•
segment
..c::
.•...•
..
<1l
.•...•
segment

-Seo
..
<1l
Cone

eo
Q
<1l
a to
Q
<1l
E Q
eo <1l
a c:
a:l
a
.2
~
<1l ell <1l ell <1l ell <1l :1:l 0::
.....l
is .....l
6 .....l
a .....l
6
(p,) (p,) (p,) (p,) (p,) (p,) (p,) (p,)
N egaprion hrevirostris* 16 2.6 15 2.3 8 1.5 11 4.7 12: 1
Gingiymostoma cirratum 17 2 16 2.5 8 1.5 10 6.2 7: 1
Dasyatis sayi 26 2 20 2 9 \.5 14 8 5:1
* From Gruber, et ai., 1963
1054 Bulletin of Marine Science (15(4)
The cones of Ginglymostoma were short and conical and measured
only 18 fl, in overall length. The outer segments were tapered and ex-
tended only to the level of the rod ellipsoid. They were lightly stained
and therefore difficult to see and photograph.
The inner segments of the cones averaged ] 0 fl, in length and 6.2 fl,
in diameter at the level of the parabaloid. The ellipsoids were heavily
stained while the parabaloids appeared empty except for a few large
granules.
Most of the nuclei of the cones of Ginglymostoma were difficult to
differentiate from those of the rods and in this way resembled those of
Negaprion. However, a significant number of the cone nuclei straddled
the outer nuclear limiting membrane and were thus easily identified. (see
Fig. 1).
A tangential section through the posterior pole of the eye is shown
in Figure 2. This photograph demonstrates clearly the difference in the
size of the inner segments of the rods and cones as well as the ratio

FIGURE 2. Tangential section of the retina of Ginglymostoma cirratum at the

level of the cone ellipsoid. Oil immersion, Mallory Triple Stain.
1965] Hamasaki & Gruber: Elasmobranch Photoreceptors 1055
of rods to cones. The ratio in the posterior pole of the eye was 7: 1. A
count of the cones from representative areas of the retina showed that
the number in the central retina exceeded that of the peripheral retina
by a ratio of 3:2.
Dasyatis sayi.-Eyes from two adult Dasyatis sayi (greatest width, 60 and
77 em) were examined, and again two types of photoreceptors were found
in the retina (Fig. 3). The cones, however, were much more numerous than
in Negaprion and Ginglymostoma and had all of the morphological features
of cones. They were short and plump and measured 23 /.f. in overall length.
The tapered outer segment did not extend to the pigment epithelium and
contributed only 9 /.f. to the overall length. The outer segments appeared to
be very fragile for many cones were noted without them. Those present
were stained only lightly which made them difficult to see and photograph.
The inner segments of the cones were thick and conical and averaged 8 /.f.

FIGURE3. Vertical Section of the retina of Dasyalis sayi. Five cones can be
seen in this section. Oil immersion; Mallory's Triple Stain; p.e.-pigment
epithelium; l.r.c.-Iayer of rods and cones; e.l.m.-external limiting membrane;
o.n.l.-outer nuclear layers; e-ellipsoid; p-parabaloid.
1056 Bulletin of Marine Science [15(4)
in diameter at the level of the parabaloid. The ellipsoids were heavily
stained while the parabaloids appeared empty except for a few large
granules.
Most of the cone nuclei were situated on the receptor side of the outer
limiting membrane and were thus located in the "myoid" portion of the
cone. The others were located immediately vitread of the outer limiting
membrane. Because of this arrangement, it was very easy to differentiate
the nuclei of the cones from those of the rods.
The rods of the Dasyatis were very similar to the rods of the other two
species examined. Thus, they were tall and cylindrical and extended into
the pigment epithelial cells. They had an overall length of 46 p.. with the
outer segment contributing 26 p.. to this. The diameter of the outer and
inner segments measured 2 p...
The inner segment of the rods consisted of the heavily stained ellipsoid
and the lighter stained parabaloid.
A tangential section, similar to that presented for Ginglymostoma, is

FIGURE 4. Tangential section of the retina of Dasyatis sayi. Because the sec-
tion was not perfectly tangential both cone ellipsoids and parabaloids can be
seen. Oil immersion, Mallory's Triple Stain.
1965] Hamasaki & Gruber: Elasmobranch Photoreceptors 1057
shown in Figure 4. Inasmuch as the plane of sectioning was not perfectly
tangential, both cone ellipsoids and parabaloids are seen. Note that the
large ellipsoids and parabaloids are surrounded by a single row of rods.
Contrary to the observation in Ginglymostoma, the number of cones in the
peripheral and central third of the retina were approximately the same. The
ratio of rods to cones in the posterior pole of the eye was 5: 1.
In both the Ginglymostoma and the Dasyatis as in the Negaprion, the
cones did not have oil droplets, and there was no evidence of double or
twin cones.
DISCUSSION

The observations of this study demonstrate clearly the presence of two

types of photoreceptors in the retina of Ginglymostoma and Dasyatis as was
the case in Negaprion. The ones designated as rods were similar in all
respects in the three species examined and possessed all of the morpho-
logical features of vertebrate rods. The cones, although slightly different in
the three species, all had small tapered outer segments and thick, plump,
inner segments. Thus the two types of receptors had the main morpho-
logical features which differentiate rods from cones (see Pedler & Tilly,
1964) .
At the light microscope level, there was little difference noted in the fine
structure of the component parts of the cones. Pedler and Tilly, after exam-
ining the receptors of diurnal and nocturnal Geckos, hypothesized that
different parts of the receptors can evolve separately and there need not be
a transmutation of the entire receptor. If this is true then it would be of
interest to study the fine structure of the cones of the elasmobranchs by
electron microscopy to see if the component parts have been modified.
The main difference in the cones of the three species, other than in size
and number, was in the location of their nuclei. Most of those of Dasyatis
were situated on the receptor side of the outer limiting membrane and
easily identified, while those of Negaprion were located deep in the outer
nuclear layer and difficult to differentiate from those of the rods. The
position of the cone nuclei in Ginglymostoma varied; some resembled those
of Negaprion in being situated deep in the outer nuclear layer, the others
straddled the outer limiting membrane and in this way resembled those of
Dasyatis.
In all three species, the ERG changed from a photopic type in the
light-adapted state to a scotopic type when the animals were dark-adapted.
However, when the dark-adaptation curve was determined using the ERG
as a measure of retinal sensitivity, a "break" in the dark-adaptation curve
was not found (see Hamasaki & Bridges, 1964). Thus, in spite of the
presence of rods and cones in the retina, there did not appear to be a
separate photopic and scotopic mechanism which determined retina
sensitivity in these three elasmobranchs.
l058 Bulletin of Marine Science [15(4)
The range of dark-adaptation was about the same for all three species
when measured by the electrophysiological method. After intense light-
adaptation the sensitivity of the retina was found to increase by 105 to 106
fold after 45 to 60 minutes in the dark. This range is comparable to that of
humans and does not reflect the differences in the rod: cone ratio found
in elasmobranchs.
SUMMARY

A histological examination of the retinas of three nurse sharks, Gingly-

mostoma cirratum, and two sting rays, Dasyatis sayi, demonstrated two
types of photoreceptors which were identified by their morphological fea-
tures as rods and cones. In the nurse shark, the rods were tall and cylin-
drical with large outer segments. The total length of the rods was 33 JL of
which 17 JL were the outer segments. The cones of the nurse shark were
short and conical and measured only 18 JL in overall length. The outer
segments were short and tapered and did not extend into the pigment
epithelial cells. Most of the nuclei of the cones were situated deep in the
outer nuclear layer and in this way resembled those of the lemon shark.
However, a significant number were found either to straddle the outer
membrane or to be located immediately vitread of the outer limiting mem-
brane. The ratio of rods to cones in the posterior pole of the eye was 7: 1.
The rods of the sting ray were similar to those of the nurse shark and
were tall and cylindrical with large outer segments. Their overall length was
46 JL, of which 26 JL were the outer segments. The cones were much more
numerous in the sting ray than in the nurse or lemon sharks. They were
short and plump with tapered outer segments, and measured only 23 JL in
overall length. Most of the cone nuclei were situated on the receptor side
of the outer limiting membrane and thus easily identified. The others were
located immediately vitread of the outer limiting membrane and were thus
similar to those of the nurse shark. The ratio of rod to cones in the
posterior pole of the eye was 5: 1.
In both the nurse shark and the sting ray, the cones were not found
to possess oil droplets.
SUMARIO
Los FOTORECEPTORES DEL TIBURON GATA, Ginglymostoma cirratum, Y
DE LA RAYA, Dasyatis sayi

Un examen histo16gico de las retinas de tres tiburones gatas, Ginglymo-

stoma cirratum, y dos rayas, Dasyatis sayi, mostraron dos tipos de fotore-
ceptores que fueron identificados par sus caracteres morfol6gicos como
bastones y conos. En el tibur6n gata los bastones eran altos y cilindricos
con grandes segmentos externos. EI largo total de los bastones fue 33fL
de las cuales 17fL eran de los segmentos externos. Los conos del tibur6n
gata fueron cortos y c6nicos y midieron s610 18fL de longitud total. Los
1965] Hamasaki & Gruber: ELasmobranch Photoreceptors 1059
segmentos externos fueron cortos y conicos y no se extend ian dentro de
las ceIulas epiteliales de pigmento. La mayorfa de los nucleos de los conos
estaban situados profundamente en la capa nuclear extern a y en esta forma
semejaban los del tiburon galano. Sin embargo, un numero significativo
fue encontrado bien cerca de la membrana limitante externa, 0 situados
inmediatamente despues de la membrana limitante externa. La proporcion
de bastones y conos en el polo posterior del ojo fue 7: 1.
Los bastones de la raya fueron similares a los del tiburon gata y fueron
altos y cillndricos con grandes segmentos externos. Su longitud total fue 46p.
de las cuales 26p. eran de segmentos externos. Los conos fueron mucho
mas numerosos en ]a raya que en los tiburones gatas 0 en los galanos.
Fueron cortos y ramos, con segmentos externos conic os y solamente
midieron 23p. de longitud total. La mayorfa de los nucleos de los conos
estaban situ ados en e] lado receptor de la membrana limitante externa y por
tanto faci]mente identificable. Los otros estaban situ ados inmediatamente
antes de la membrana 1imitante externa y eran por tanto similares a los del
tiburon gata. La proporcion de bastones y conos en el polo posterior de]
ojo fue 5:1.
Tanto en e] tiburon gata como en la raya, no se encontr6 que los conos
tuvieran gotas de aceite.

LITERATURE CrrED
GRUBER, S. H., D. 1. HAMASAKI, AND C. D. B. BRIDGES
1963. Cones in the retina of the lemon shark (Negaprion brevirostris)
Vision Res., 3: 397-399.
HAMASAKI, D. I. AND C. D. B. BRIDGES
1964. Properties of the fish e.r.g. (Abstract) 4th Internal. Photobiology
Congress. Oxford, England.
PEDLER, C. AND RITA TILLY
1964. The nature of the gecko visual cell. A light and electron microscopic
study. Vision Res., 4: 499-510.
ROCHON-DuVIGNEAUD, A.
1943. Les Yeux et la Vision des Vertebn~s. pp. iii+7J9, figs. 1-500,
Masson, Paris.
WALLS, G.L.
1942. The vertebrate eye and its adaptive radiation. pp. vii+ 785, pI. 1,
197 figs. Cranbrook Press, Michigan.