Accepted Article

DR. KEVIN K. HAUSSLER (Orcid ID : 0000-0003-3860-0282)

Article type : General Article

Editorial ref. code: EVJ-GA-19-331.R3

Contrast therapy: Tissue heating and cooling properties within the equine distal limb

Kevin K. Haussler*1, Shana R. Wilde1, Michael S. Davis2, Ann M. Hess3 and C. Wayne McIlwraith1

1Gail Homes Equine Orthopedic Research Center, College of Veterinary Medicine and Biomedical
Sciences, Colorado State University, Fort Collins, Colorado, USA;
2Department of Physiological Sciences, College of Veterinary Medicine, Oklahoma State University,
Stillwater, Oklahoma, USA and
3Department of Statistics, Colorado State University, Fort Collins, Colorado, USA.

*Corresponding author email: Kevin.Haussler@colostate.edu

Keywords: horse; contrast therapy; cryotherapy; tissue heating; distal limb

Running title: Contrast therapy applied to equine distal limb

Summary
Background: Rehabilitation of tendon injuries in horses often involves cryotherapy to reduce
inflammation and occasionally tissue heating to increase collagen extensibility. The application of
alternating cold and hot (i.e. contrast therapy) is widely used in human physical therapy; however, its utility
in equine rehabilitation is largely unknown.
Objectives: The objectives of this study were to 1) assess if the equipment could achieve therapeutic tissue
temperatures (< 15C and > 40C) at different tissue depths relative to the digital flexor tendons and 2)
evaluate the time-temperature profiles during serial heating and cooling cycles using a contrast therapy
device.
Study design: In vivo experiment.

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differences between this version and the Version of Record. Please cite this article as doi:
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 Methods: In 4 adult horses with normal forelimb digital flexor tendons, fine-wire temperature probes were
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placed superficially on the skin and implanted subcutaneously, deep to the superficial digital flexor tendon
(SDFT), and deep to the deep digital flexor tendon (DDFT). Temperatures were recorded over three
complete thermal (hot-cold) cycles. Minimum and maximum temperatures were recorded and the rate of
temperature changes and the areas underneath the time-temperature curves (i.e. thermal load) were
calculated.
Results: Minimum and maximum tissue temperatures (C) included: superficial skin [12.6 ± 1.0; 42.4 ±
2.4], subcutaneous tissues [14.1 ± 0.8; 42.3 ± 2.2], deep to the SDFT [15.6 ± 0.8; 41.7 ± 2.6], and deep to
DDFT [25.1 ± 2.0; 38.0 ± 3.5]. An initial rapid rate of tissue temperature change between 3.2 to 4.3C/min
occurred within tissues to the depth of the DDFT. Tissue thermal loads during heating ranged from 255 to
607C*sec and from 309 to 780C*sec during tissue cooling, with the lower values noted deep to the
DDFT.
Main limitations: Unknown clinical efficacy in diseased tissues.
Conclusions: The applied contrast therapy was consistently able to induce cooling and heating of tissues
to the depth of the DDFT.

Introduction
Topical heating is primarily used for increasing soft tissue compliance and flexibility while reducing pain
and muscle hypertonicity [1,2]. Most superficial heating modalities have limited ability to heat deep tissues
without an increased risk for inducing thermal injuries to the overlying skin [3]. Cryotherapy or tissue
cooling is most often used to reduce inflammation and pain following an acute injury or post-operatively
[4]. Cryotherapy of the equine distal limb has been used with varying degrees of efficacy using cold packs,
sleeves with circulating cold media, and ice water immersion [5,6]. Comparison of wet-interface
(immersion, ice boots) versus dry-interface (pneumatic sleeve, cold packs) showed that cold water
immersion and ice boot application achieve superior cooling of tissues in the foot, while a pneumatic
device was the only dry-interface technique to achieve tissue temperatures within the therapeutic range (4
to 7C) [3,5]. Reducing tissue temperatures to 10-15C is considered necessary to maximise the
physiological effects of vasoconstriction, pain relief and decreased metabolic rates [7]. However, the
haemodynamic effects of cold therapy often prevent effective cooling of soft tissue structures deeper than
2-3 cm, regardless of duration of the applied cold therapy [8].

Contrast therapy is characterised by repeated, alternating applications of both cold and heat to aid in post-
exercise recovery and to treat acute soft tissues injuries with the aim of achieving increased blood
circulation through cyclic vasodilation and vasoconstriction [9]. In human studies, there is agreement that

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 the temperatures applied at the skin should be 10-15C for cryotherapy and 38-43C for heating modalities
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[9,10]. In horses, the ability of a contrast therapy device to deliver target hot and cold temperatures is
unknown. The objective of this experimental study was to investigate if a dry-interface contrast therapy
unit could consistently and effectively induce therapeutic tissue temperatures (< 15C and > 40C) relative
to the superficial and deep digital flexor tendons within the equine distal limb. A secondary aim was to
evaluate any effects due to the order of applied treatment (i.e. heat-first or cold-first).

Materials and Methods

Horses
Four mixed-breed, adult horses [mean ± s.d. age = 3.8 ± 1.0 years; 3 female and 1 male; weight = 460 ± 36
kgs] with grossly palpable normal forelimb digital flexor tendons and no concurrent lameness were used in
this study. A single, randomised forelimb within each horse was instrumented for each data collection
session. Approximately 7-10 days later, the opposite forelimb was then instrumented and used for data
collection.

Thermistor Placement
Horses were placed in a portable stock for restraint during data collection using intravenous sedation
(detomidine hydrochloride 0.01 mg/kg bwt).a To facilitate thermistor placement, local perineural
anaesthesia of the lateral and medial palmar and palmar metacarpal nerves at the level of the proximal third
of the metacarpus was completed using 3 ml of 2% mepivacaine hydrochloride (Carbocaine-V)a per site.
Hair was clipped circumferentially from the metacarpal region and the skin overlying the lateral aspect of
the digital flexor tendons was disinfected. Three sterilised temperature probes (Model IT-18)b were
threaded through 18-gauge needles and placed 1) subcutaneously, 2) deep to the superficial digital flexor
tendon (SDFT), and 3) deep to the deep digital flexor tendon (DDFT) with ultrasound-guidance (Fig 1).
The thermistors were held in place with cyanoacrylate glue applied to the skin at the insertion site and
anchored with medical tape to the forelimb. An additional thermistor (Model MT-4)b was applied topically
on the skin along the dorsal metacarpus to monitor superficial skin temperatures within the instrumented
limb. A layer of plastic wrap was placed over the metacarpal region to protect the thermistor insertion sites
from the overlying circulating sleeve used to apply the contrast therapy. The circulating sleeve was then
placed over the metacarpal region, and a cotton standing wrap and self-adherent cohesive bandaging tape
was applied to help provide compression and temperature retention. A surcingle was used for securing
tubing and wire leads from the forelimb. Two additional thermistors (Model MT-4)b were placed 1)
topically in the wither region underneath the surcingle to monitor surface body temperatures on haired skin
and 2) free within the testing area to monitor ambient temperatures. Lateral radiographs of the distal
metacarpus were taken to assess relative placement of all implanted thermistors (Fig 1). Transverse

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 ultrasound images of the digital flexor tendons were also performed to confirm placement of the
Accepted Article
thermistors relative to the superficial and deep digital flexor tendons.

Application of Contrast Therapy

Tubing that extended from the circulating sleeve was attached to the external contrast therapy unit
(Vorteq).c The inflow and outflow tubing circulated monitored heated or cooled fluid through the sleeve to
provide tissue heating or cooling to the metacarpal region. The contrast therapy unit allowed precise
delivery of programable and cyclic temperatures over set times. The initial application of hot or cold was
randomly assigned either to the left or right limb of the horse to assess the effects of the order of the applied
tissue heating or cooling. Each temperature cycle consisted of 15 minutes of applied heat at 48C and 15
minutes of cold therapy at 7C, as measured at the contrast therapy device. Three complete thermal
(hot:cold) cycles were then applied to each limb within each session for an overall total of approximately 2
hours per session (Fig 2). Temperature readings from the thermistors were collected at 15-second intervals
using data acquisition hardware (Thermes)b and software (DASYLab)b installed on a laptop computer.
Based upon pilot work, a circulating fan was used to cool the contrast therapy unit to help increase its
cooling efficiency when applied during days with high ambient temperatures (>30C) within the closed
testing area. After each data collection session, the thermistors were removed, and light bandages were
applied for 3 days to help reduce the risk for soft tissue swelling and infection. Nonsteroidal anti-
inflammatories and antibiotics were given for 3 days. Approximately 7-10 days later, the above procedures
were repeated on the contralateral forelimb within each horse.

Data Processing
Peak temperatures (i.e. maximum and minimum) during the tissue heating and cooling phases were
measured to determine if the contrast therapy device could effectively heat tissues to > 40C and cool tissue
temperatures to < 15C within the equine distal limb (Fig 3). The time to reach target tissue temperatures at
different depths were determined across heating and cooling cycles to identify the duration of treatment
needed to reach the respective target temperatures. The rate of tissue heating and cooling (i.e. slope of time-
temperature curves) was calculated at the initial and final phases of temperature changes to evaluate the
efficacy of the contrast therapy device in altering tissue temperatures during cyclic tissue heating and
cooling (Fig 3). The initial rate of tissue temperature change was calculated within the linear region
immediately following the initiation of a specific thermal cycle (heating or cooling). The final rate of tissue
temperature change was measured within the linear region immediately preceding the switch to the
alternate thermal cycle. The area under the heating and cooling curves were calculated using integration
and defined as a measure of tissue thermal load (Fig 3). Changes in the above parameters were measured

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 for each hot:cold cycle (for example, A vs. A’ vs. A”, Fig 2) to determine if there were residual thermal
Accepted Article
effects due to repeated hot-cold cycles and to evaluate if there was a significant thermal effect due to the
order of the applied treatment (i.e. heat-first or cold-first). Fluctuations in the minimum and maximum
temperatures were deemed clinically significant if > 2C between three hot-cold cycles within a treatment
session.

Temperature differences were calculated between the 1) heated or cooled fluid delivered by the contrast
therapy device (i.e. supply), 2) skin temperature, and 3) return fluid temperatures measured within the
device to estimate thermal losses within the tubing and compression sleeve and due to thermal conduction
within the distal limb during tissue heating and cooling phases.

Data Analysis
Descriptive statistics were used to report minimum and maximum tissue temperatures, rates of tissue
heating and cooling, and areas under the heating and cooling curves. Formal statistical tests were used to
assess the effect of the order of the applied treatment (i.e. heat-first or cold-first). A mixed model was fit
for each response variable at each of the four tissue depths separately. To assess the efficiency of the
thermal delivery by the device, additional tests were completed that included the superficial skin and fluid
supply and return temperatures as response variables. Initial hot or cold application was included as a fixed
effect. Horse was included as a random effect to account for repeated measures. We note that response
variables are likely correlated, but univariate analysis was used. Visual inspection of residual diagnostic
plots was used to evaluate model assumptions of normality and equal variance. All tests assumed a 2-sided
alternative hypothesis (comparing heat-first versus cold-first) and P<0.05 was considered statistically
significant. All analyses were performed using JMP statistical software version 15.0.d

Results
Peak Temperatures
The pooled minimum and maximum tissue temperatures (C) at the different tissue depths were: superficial
skin [12.6 ± 1.0; 42.4 ± 2.4], subcutaneous tissues [14.1 ± 0.8; 42.3 ± 2.2], deep to the SDFT [15.6 ± 0.8;
41.7 ± 2.6], and deep to DDFT [25.1 ± 2.0; 38.0 ± 3.5] (Fig 4).

Target Tissue Temperatures

The skin, subcutaneous and SDFT tissues consistently reached target tissue temperatures of > 40C and <
15C (Fig 4). Target tissue temperatures at the skin were reached within 11 minutes, independent of
whether heat or cold was initially applied. Target temperatures at the skin were reached within 9.5 minutes

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 when heat was applied first but required up to 12.5 minutes when cold was initially applied. In the majority
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of limbs, tissues deep to the DDFT did not reach target tissue temperatures during the 15-minute heating or
cooling cycles (Fig 4).

Rate of Tissue Temperature Changes

The initial rate of temperature change occurred within the first 5 minutes of hot or cold application and was
most rapid within the subcutaneous tissue location and was significantly slower for tissues deep to the
DDFT (Fig 5). The initial rapid rate of tissue temperature change occurred within the range of 16C to
36C at the skin surface, 19C to 36C at the subcutis and deep to the SDFT, and 27C to 34C deep to the
DDFT (Example shown in Fig 2). During initial tissue cooling, the linear region of the time-temperature
curve extended well below the baseline tissue temperatures (e.g. 20C versus 30C baseline temperature).
Conversely, during the initial phase of tissue heating the linear region typically did not extend above
baseline tissue temperatures. The final rate of temperature change was consistent across tissue depths (Fig
5); however, the onset occurred earlier during cooling (~ 4.5 minutes), compared to the heating phase (~ 6
minutes).

Tissue Thermal Load

Tissue cooling occurred more rapidly and maintained a target tissue temperature for longer periods of time,
compared to tissue heating (Fig 6).

Order of Applied Treatment

The order of the applied treatment (i.e. heat first or cold first) was not statistically significant (all p > 0.2)
or clinically relevant (i.e. differences >2C) for the minimum or maximum temperatures measured across
the three hot:cold cycles within each treatment session. Therefore, all response variables were averaged
across the three hot:cold cycles within a treatment session. For maximum skin temperatures, if heat was
initially applied, then the temperatures for the first versus last cycle decreased by 2C (e.g. 43 to 41C);
whereas, if cold was initially applied, then the temperatures for the first versus last cycle increased 2C
(e.g. 41 to 43C). For the minimum tissue temperatures there was increased variability at the different
tissue depths, but if heat was initially applied, then the subcutis temperatures for the first versus last cycle
decreased by about 1C (e.g. 14 to 13C). Whereas, if cold was initially applied, then the subcutis
temperatures for the first versus last cycle increased about 2C (e.g. 12 to 14C). There were significant
differences in the initial rate of temperature change during both heat-first (p < 0.01) and cold-first (p <
0.01) conditions. Applying heat first reduced the initial rate of tissue heating due to relatively smaller

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 temperature differences needed to heat tissues beginning at body temperature (i.e. from ~33C to 44C),
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compared to cooling tissues first and then applying heat (i.e. from ~13C to 44C).

Efficiency of Temperature Delivery

With no tissue contact, there was a 0.3C temperature loss through the tubing and attached circulating
sleeve during both the heating and cooling phases. When the sleeve was in contact with the distal limb, the
supply temperature was 47.7  0.2C across all the heating phases and was 7.3  0.7C across all cold
phases. The order of the applied treatment (i.e. heat-first or cold-first) only had a significant effect on the
return temperatures measured during tissue cooling (Table 1). When comparing differences between the
supply, skin and return temperatures, the majority of thermal change occurred due to tissue interaction (~
5C) versus equipment or environmental losses (~ 0.5C).

Discussion
The contrast therapy device used in this study was able to consistently heat and cool superficial tissues
within the distal limb to target temperatures as hypothesised. However, tissues deep to the DDFT did not
reach target temperatures and the measured temperatures were often quite variable between horses and hot-
cold cycles. The contrast therapy device was able to repeatably cool and heat tissues over the three hot:cold
cycles lasting approximately 2 hours. This is a significant improvement over other forms of topical heat or
cold therapies which often produce wide-fluctuations in tissue temperatures depending on the type and
application [11]. The ability to measure in real-time the temperature of the fluid media being delivered to
the tissues is critical from a therapeutic perspective and has important safety implications that are not
possible with any other forms of cryotherapy or heating modalities [3,6]. Dry-interface pads also limit skin
exposure to water and the need to manage ice-melt. Desmitis, cellulitis, tissue necrosis and distal limb
oedema have been reported with ice-water contact, which can be mitigated with the application of dry-
interface sleeves [6].

The rapid, initial rate of tissue temperature change measured within the superficial tissues occurred within
the range of approximately 18C to 36C. Given the average baseline skin temperature of 32C in our
study, the above range reflects a 4C increase and 14C decrease in tissue temperatures, which implies that
tissues generally resist heating more than cooling. This is also confirmed by measures of increased tissue
thermal loads during the cooling versus heating phases. Tissue heating is associated with a rapid
vasodilation of local vessels and subsequent increase in blood flow [1,9]. Initial temperature changes
measured immediately after peak heating or cooling but prior to reaching baseline tissue temperatures were
likely due to reversal of the applied contrast temperatures combined with tissue perfusion effects; whereas,

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 continued temperature changes above (or below) baseline tissue temperatures were likely due to the
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contrast therapy unit driving the temperature change by itself. However, there were no significant
differences in the rate of temperature change measured before or after the baseline tissue temperatures had
been reached. Therefore, the initial rate of tissue temperature changes was likely due to applied hot or cold
and less due to tissue perfusion effects. It is likely that the initial rate of temperature change induced by the
contrast therapy unit was too rapid and overrode any underlying tissue perfusion effects.

The discrepancy in tissue responses to applied hot or cold may indicate that physiologically tissues have
more effective methods for removing excess heat to protect against heat stress, compared to tissue cooling.
Vasodilation and increased blood flow, especially in highly vascular tissues (e.g. skin, viscera) and areas
with large muscle mass (e.g. proximal limbs and epaxial regions), seem to be very effective mechanisms
for preventing tissue overheating [12]. The medial and lateral palmar vessels located adjacent to the DDFT
likely provided a countercurrent temperature mechanism that played a role in the inability to consistently
heat or cool tissues deep to the DDFT, compared to the more superficial tissues that lacked similar
vasculature. An alternative explanation was that the applied thermal stimulus was not able to penetrate the
deeper tissues; however, this was unlikely due to the circumferential application of the hot and cold. The tip
of the thermistor placed deep to the DDFT was likely an equidistance from the palmar surface and the
medial or lateral aspects of the metacarpus.

A slower, final rate of temperature change was noted at temperatures > 36C and < 18C and had a more
defined onset during tissue cooling, compared to tissue heating. This implies that once tissue is cooled
below approximately 18C then additional protective mechanisms beyond vasoconstriction are engaged to
help slow tissue cooling and prevent thermal injury [13]. The shunting of blood flow to deeper tissues is
responsible for rapid cooling of superficial tissues and the delayed cooling of deeper tissues [10,14].
Similar protective mechanisms likely occur during tissue heating; however, the onset is more gradual and
less noticeable [15]. It is unknown if the final rate of tissue heating or cooling is due to solely to
haemodynamic effects, which obviously have physiologic limits in regulating tissue heating or cooling
[16].

Interestingly, there were measurable, albeit inconsistent, differences in the order of initial heating or
cooling of tissues that disappeared once the device had completed a full hot:cold cycle. It is not clear if
these changes were due to differences in temperature delivery by the contrast therapy device or due to
actual physiologic tissue responses. If heat was applied first, then the supply temperature from the contrast
therapy device during the cold phase was ~8C, compared to if cold was applied first (~7C). This implies

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 that the contrast therapy unit was not able to deliver cold independent of heat delivery, which may have
Accepted Article
influenced peak temperature measures over the three hot:cold cycles. The contrast therapy unit did have
challenges in delivering and maintaining defined cooling thresholds when used in a hot ambient
environment (> 30C). Adding circulating fans did aid the contrast therapy devices’ ability to produce the
desired cold output. However, it was judged that the 1-2C variability noted over repeated application was
not biologically or clinically relevant, especially given the poor consistency of heat or cold delivery
provided by other topical thermal applications (e.g. cold packs) [3,15]. Although, the time to reach
therapeutic tissue heating temperatures varied by 3 minutes when heat was applied first versus when cold
was initially applied, which may have clinical implications from an effective treatment time perspective.

Our primary goal was to assess if this contrast therapy device could safely and effectively induce target
temperatures (< 15C and > 40C) to tissues around the digital flexor tendons. Across horses, the lowest
skin temperature recorded was 9C and the highest was 44C. Literature on skin thermal injury thresholds
suggest that healthy, intact tissues can tolerate heating up to 50C, which was not reached at the skin
surface in any horse at any time point in our project [17]. Our results for tissue responses to topical heat
were comparable to prior studies [3,15]. The contrast therapy unit used in this study was able to induce
therapeutic tissue heating within 5-10 minutes, depending on tissue depth. Similarly, therapeutic tissue
cooling could be induced within 10 minutes, depending on tissue depth, which is comparable to ice water
immersion [3,18]. Although the data appear to show that contrast therapy using this specific device and
treatment protocol did not consistently achieve target temperatures in the DDFT, our data may
underestimate the temperature profiles actually achieved during this study. Our thermistors were placed in
the tissues surrounding the tendons, rather than within the tendons themselves, and it is likely that the
temperatures within the tendons themselves were the average of the measurements from the two
thermistors bracketing the tissue of interest (subcutis and SDFT for the superficial digital flexor tendon,
and SDFT and DDFT for the deep digital flexor tendon). With this inference, we can propose that target
temperatures were consistently achieved using both heating and cooling contrast therapy in the superficial
digital flexor tendon, and target temperature was likely achieved during heating in the deep digital flexor
tendon, but not during tissue cooling.

Based on pilot data, we selected 15-minute heating and cooling cycles to limit the duration of treatment and
time required to collect data from 3 complete hot:cold cycles. From the time-temperature graphs, it appears
that neither tissue heating or cooling responses had plateaued completely so the treatment times could have
been lengthened to maximise treatment effects [15]. Longer treatment times would have also likely
produced more consistent results and therapeutic temperatures in tissues deep to the DDFT.

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Accepted Article
We chose to apply consistent and equal treatment times for both hot and cold phases, despite some clinical
applications that use hot:cold ratios of 3:1 or 4:1 [10]. Other treatment protocols used in humans involve
alternating heat at 38-43C for 2-5 minutes and cold at 4-14C for 2-5 minutes over 2 to 9 thermal cycles.
Obviously, these short treatment times will not induce therapeutic tissue temperatures; however, additional
neurologic or psychological mechanisms of action are likely activated with these rapid shifts in temperature
[19]. It is likely that altering the duration of hot relative to cold would produce variable tissue responses,
which might be more favourable for treating select disease or injury conditions. Investigating the optimal
dosage parameters and timing relative to the phase of tissue healing, injury severity, and affected tissues or
location is needed in future studies [1].

The dry-interface contrast therapy used in this study was a safe and effective method for delivery of known
heating and cooling of tissues. These results help define the physiologic responses of cyclic tissue heating
and cooling at different tissue depths within the equine distal limb. Future work is needed to assess the
clinical applications of contrast therapy for pain and specific disease conditions in horses.

Authors’ declarations of interest

No competing interests have been declared.

Ethical animal research

This project was approved by the Institutional Animal Care and Use Committee, Colorado State University.

Owner informed consent

Not applicable.

Data accessibility statement

The datasets generated for use in the current study are available from the corresponding author on
reasonable request.

Source of funding
Research funding was provided by the Equine Orthopedic Research Fund, Colorado State University.
Student support provided by a USDA Fellowship - USDA-NIFA Animal Health & Disease Research
Program Funding.

Acknowledgements

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 Cascade Wellness donated the equipment used in this study. The authors thank Dr Katie Seabaugh and
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Jennifer Daniels for assisting with instrumentation and horse management.

Authorship
K. Haussler and S. Wilde participated in the study design, study execution and data collection and analysis.
All authors participated in data interpretation, manuscript preparation, and final approval of the manuscript.

Manufacturers’ addresses
aZoetis Inc., Parsippany, New Jersey, USA.
bPhysitemp Instruments Inc., Clifton, New Jersey, USA.
cCascade Wellness Technologies, Inc., Sunriver, Oregon, USA.
dSAS Institute, Cary, North Carolina, USA.

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 Figure legends
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Fig 1: Thermistor placement and implantation a) grossly and b) radiographic confirmation of thermistor
placement at the four instrumented tissue sites.

Fig 2: Representative temperature fluctuations at the different tissue depths over three hot-cold cycles
within a single forelimb. Additional temperature measurements included the skin at a control site (withers)
and within the ambient environment. Illustrated changes in peak tissue cooling (A, A’, A’’) and heating (B,
B’, B’’) over three hot:cold cycles.

Fig 3: Illustration of peak heating (A) and cooling (B) relative to the target tissue temperatures of > 40°C
(red line) and < 15°C (blue line) during a single hot-cold cycle. Sites of measured initial and final tissue
temperature change (green bars) and measured area under the time-temperature curves (tissue thermal load)
are also noted.

Fig 4: Pooled minimum and maximum temperatures (mean; standard deviation) relative to target
temperatures (> 40°C and < 15°C) across the different tissue locations.

Fig 5: Initial a) and b) final rates of cooling and heating (mean; standard deviation) at the different tissue
depths.

Fig 6: Tissue thermal load (mean; standard deviation) measured during heating and cooling phases at
different tissue depths.

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 Tables
Accepted Article
Table 1: Supply and return fluid temperatures (C) recorded within the contrast therapy device compared
to temperatures measured at the superficial skin during tissue heating and cooling phases with either heat or
cold applied initially (mean; standard deviation). a,b Different superscripts within a row indicate significant
differences depending on if heat or cold was initially applied.

Tissue Heating Tissue Cooling

Sites Heat First Cold First Heat First Cold First
Supply Temperatures 47.8  0.3 47.6  0.1 7.9  0.3 6.8  1.1
Skin Temperatures 42.9  0.5a 42.0  0.6b 13.2  0.8 12.0  0.9
Return Temperatures 42.3  0.3 41.6  0.6 13.6  0.4a 11.6  1.4b
Site Differences
Supply-Skin -4.9  0.4a -5.6  0.7b 5.3  0.6 5.1  0.6
Skin-Return -0.6  0.6 -0.4  0.9 0.4  0.5 -0.4  0.6
Supply-Return -5.5  0.5 -6.0  0.5 5.8  0.3 4.7  0.4

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