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Core and Surface Temperature Modification During Road Transport and Physical Exercise in Horse After Acupuncture Needle Stimulation
Journal of Equine Veterinary Science 55 (2017) 84–89
Contents lists available at ScienceDirect
Journal of Equine Veterinary Science journal homepage: www.j-evs.com
Original Research
Core and Surface Temperature Modification During Road Transport and Physical Exercise in Horse After Acupuncture Needle Stimulation Maria Rizzo a, Francesca Arfuso a, Elisabetta Giudice b, Francesco Abbate a, Francesco Longo c, Giuseppe Piccione a, * a
Department of Veterinary Sciences, University of Messina, Polo Universitario dell’Annunziata, Messina, Italy Department of Chemical, Biological, Pharmaceutical and Environmental Sciences, University of Messina, Messina, Italy c Veterinary Practitioner, Bologna, Italy b
a r t i c l e i n f o
a b s t r a c t
Article history: Received 7 February 2017 Received in revised form 30 March 2017 Accepted 30 March 2017 Available online 18 April 2017
In this study, the effect of acupuncture on skin temperature (TSKIN), including six body regions (neck, shoulder, ribs, flank, internal thigh, and back), rectal temperature (TRECTAL), serum levels of total proteins, hematocrit, magnesium, calcium (Ca), phosphorus, and chloride was evaluated in five Thoroughbred horses. Horses competed in two official races. For each race, animals were transported from their stables to the racetrack. Horses transported and competed in the first race represent the control group. Two weeks later, the same horses competed in the second race. Before road transport, they were treated with acupuncture. From animals, the TSKIN, TRECTAL, and blood samples were collected at rest; after unloaded, 30 minutes after unloaded, at rest in the transit box, at the end of the race, and 30 minutes after the race. A significant effect of transport and exercise was found on all studied parameters (P < .01), except for Ca and TSKIN of neck (P > .05), in both groups. Transported and exercised horses subjected to acupuncture treatment showed statistically significant higher values of TRECTAL and TSKIN of flank (P < .01). Thoroughbred horses display a classic hemodynamic response to transport and exercise; acupuncture affects body temperature and skin temperature of flank region in both transported and exercised horses suggesting a controlling effect of acupuncture on thermoregulatory function, probably, by decreasing the activities of sympathetic function and manifested as vasodilatation, arterial blood flow, and skin microcirculation increase. Ó 2017 Elsevier Inc. All rights reserved.
Keywords: Acupuncture Body temperature Exercise Horse Road transport
1. Introduction Transport and physical exercise represent stressful stimuli which can lead to homeostasis disruption with direct impact on animal health status and physical
Conflict of interest: The authors disclaim any financial support or relationships that may pose conflict of interest. * Corresponding author at: Giuseppe Piccione, Department of Veterinary Sciences, University of Messina, Polo Universitario dell’Annunziata, 98168, Messina, Italy. E-mail address: [email protected] (G. Piccione). 0737-0806/$ – see front matter Ó 2017 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.jevs.2017.03.224
performance of the athletic horse [1–3]. Athlete horses are used to travel before competitions, but, after transport, some of them show lower performance than usual [4]. To limit health problems related to transport and exercise stress, it is important to evaluate the health status of the horses before and after the physical effort, to optimize the environmental conditions inside the truck or to provide them with electrolytes and antioxidants. The evaluation of body temperature represents a valuable tool to monitor the physiologic status, welfare, and the stress response of animals. Monitoring horse surface temperature allows the evaluation of function of individual parts of the body. The
M. Rizzo et al. / Journal of Equine Veterinary Science 55 (2017) 84–89
assessment of horse surface temperature changes resulting from transport and/or exercise could allow detecting inflamed areas caused by stress of musculoskeletal system that could account for a decreased welfare and physical performance [5]. During exercise, about 20% of the metabolism in the muscle cells is used for work and the remaining 80% becomes heat [6]. Similarity to human, skin is the only heat dissipation way of the equine body; when the body energy metabolism and heat production increases, heat dissipation through the skin increases, too, and so the surface temperature rises [7]. Heat removal rate is generally determined by the heat transfer rate from the core to the surface of the body through blood vessels. In particular, vasodilation allows more blood to flow through the arteriole, thus venting the heat by convection and conduction through the superficial capillaries, if vasodilatation is not sufficient sweating starts [8]. Sweating can lead to fluid and electrolyte losses that if uncompensated, can lead to thermoregulatory instability, hypovolemia, imbalance in electrolyte serum concentration, and when the deficit becomes serious, horses can show clinical signs of dehydration compromising the physical performance [1,9]. Sweat loss of electrolyte, in particular, causes the onset of peripheral fatigue and weakness [10]. It has been demonstrated that acupuncture treatment leads to alleviation of muscle tension, improvement of local blood flow, increase of pain threshold, and modulation of autonomic nervous system [11,12]. Acupuncture stimulation induces alterations in microcirculation, including the increase in the skin and muscle blood flow [13], and it lead to the release of biological factors, such as nitric oxide [14,15] and calcitonin gene-related factor [16], which play important roles in the modulation of vasodilation [17]. It has been well established that acupuncture can influence skin temperature and blood flow in human [18,19]. However, to the best authors’ knowledge, no studies have been currently published on the influence of acupuncture treatment on thermoregulation and hydration status of equine species. Therefore, the aim of the present study was to evaluate how the values of rectal and skin temperatures as well as hydration indices change following acupuncture stimulation in Thoroughbred horses before and after road transport and physical exercise. 2. Materials and Methods 2.1. Animals and Study Design All treatments, housing and animal care reported below were carried out in accordance with the standards recommended by the EU Directive 2010/63/EU for animal experiments. Five clinically healthy and regularly trained Thoroughbred horses (five geldings, mean age 4 1 year; mean body weight 437 15 kg) were used with the informed consent of the owners. Animals were stabled in individual boxes (3.5 3.5 m) at the same training center located in Sicily, Italy (38 000 4900 N, 15 2501800 E, 80 m above sea level) under natural photoperiod (sunrise at 6:11 AM, sunset at 5:13 PM; mean temperature 23 C, relative humidity 70%). Horses were fed, twice a day (7 AM and 5 PM),
85
a total food amount of about 2.5% of horse body weight (forage-to-concentrate ratio 70:30), and water was available ad libitum. The horses competed in two official 1,300meter races at “Mediterraneo” racetrack (Siracusa, Sicily, Italy). For each race, the five horses were transported from their stables to the racetrack in a 5-horse truck. Each animal traveled, tethered with a 50-cm rope on each side of the halter, in an individual tie box (length 2.3 m; width 0.85 m), giving a total space of about 2 m2 and made two journeys of 145 km. Each journey was made 2 weeks after the previous one. The driver and the route were always the same. Thermal and hygrometric records were carried out inside the box for the whole study by means of a data logger (Gemini, UK). The same instrument was placed approximately 5 cm from the center of the ceiling in the van during transport in order to measure the ambient thermal and hygrometric records inside the truck throughout the transport. The temperature in the box ranged between 24 C and 16 C (maximum/minimum), whereas the relative humidity ranged between 25 C and 67%. Temperatures inside the truck ranged between 24 C and 21.5 C (maximum/minimum). The relative humidity inside the truck ranged between 68% and 75%. Horses transported and competed in the first official race represent the control group (CG). Two weeks later, the same horses competed in the second official race. Before road transport, they were subjected to acupuncture treatment (acupuncture group, AG) and were loaded after 30 minutes onto the truck calmly and without the use of force. Acupuncture was performed once a day for 30 minutes across all animals. A stainless-steel needle (diameter 0.25 mm, length 30 mm, Dongbang Acupuncture Inc, Qingdao, China) was inserted vertically into a depth of 2–3 cm, and stimulation was produced by bidirectional twisting of needles, as described in the previous studies [20,21]. In particular, Table 1 shows the name and location of the selected acupoints received from acupuncture group (AG). For acupuncture stimulation, stainless needles were inserted into the left or right side of selected acupoints. The locations of acupoints were found according to the anatomical structures. 2.2. Body Temperature Measurement From each animal, rectal and skin temperatures were measured by the same operator at rest in their box at 8:30 AM; after being unloaded from the vehicle and housed in the transit boxes, 30 minutes after unloaded, at rest in the transit box, at the end of the race, and 30 minutes after the race. Rectal temperature (TRECTAL), taken as representative of body temperature, was measured by a means of a digital thermometer (HI92704, Hanna Instruments Bedfordshire, UK), inserted 15 cm in the rectum. Skin temperature (TSKIN) was measured at six locations (neck, shoulder, ribs, flank, internal thigh, and back) with an infrared thermometer (Testo 826-T1) having a sensitivity of 0.2 C. 2.3. Blood Sampling and Laboratory Analysis From each animal, blood samples were collected by the same operator at the same time points of the body
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M. Rizzo et al. / Journal of Equine Veterinary Science 55 (2017) 84–89
Table 1 Name and location of the acupoints selected for the study. Chinese Name
Other Name Location
Ming-men (life gate)
GV 4
Da-zhui (big vertebra)
GV 14
Dan-zhong (chest center)
CV 17
Qian-chan-wan (thoracic fetlock)
SI 3
Shen-mai (extending vessel) Shang-liao (upper bone hole) Ci-liao (second bone hole) Zhong-liao (central bone hole) Xia-liao (lower bone hole) Huang-men (vitals gate) Fei-pan (lung hugging) Hua Tuo Jia Ji (Hua Tuo’s paravertebral points)
BL BL BL BL BL BL
a
62 31 32 33 34 52
In the depression along the dorsal midline at the intervertebral space between L2 and L3 In the depression along the dorsal midline at the cervicothoracic intervertebral space (C7–T1) On the ventral midline, at the level of the caudal border of the elbow, or at the level of fourth intercostal space In a depression on the caudolateral border of the cannon bone (third metacarpal), distal to the end of the lateral splint bone (fourth metacarpal), proximal to the fetlock, over the lateral palmar digital vein In a depression distal to the lateral malleolus First sacral intervertebral space (S1–S2), 1.5 cuna lateral to the dorsal midline Second sacral intervertebral space (S2–S3), 1.5 cuna lateral to the dorsal midline Third sacral intervertebral space (S3–S4), 1.5 cuna lateral to the dorsal midline Fourth sacral intervertebral space (S4–S5), 1.5 cuna lateral to the dorsal midline At the level of the lower border of the spinous process of L1, 3 cuna lateral to the GV-5. On the caudal border of the scapula, 1/3 of the distance from dorsal to the ventral 0.5–1 cuna lateral to the depressions below the spinous processes of the L2–L6.
The cun is a relative or proportional measurement unit, which is used to locate the Jing-Luo and acupuncture points.
temperature recording. Blood samples were collected by jugular venipuncture into 2-mL vacutainer tubes containing ethylenediaminetetraacetic acid and into 8-mL vacutainer tubes with cloth activator (Terumo Co, Tokyo, Japan). Immediately after collection, blood samples were placed in refrigerated bags and transported to the laboratory for the analysis. ethylenediaminetetraacetic acid whole blood samples were processed in the laboratory within 2 hours for the evaluation of hematocrit (Hct) by means of an automated hematology analyzer (HeCo Vet C, SEAC, Florence, Italy). Blood samples collected into tubes containing clot activator were centrifuged at 1,300g for 10 minutes, and the obtained sera were analyzed within 2 hours to estimate the concentration of total proteins (TPs), magnesium (Mg), calcium (Ca), phosphorus (P), and chloride (Cl) using commercially available kits by means of an automated analyzer UV Spectrophotometer (model Slim SEAC, Florence, Italy). 2.4. Statistical Analysis Data, expressed as mean values standard deviation, were tested for normality using the Shapiro–Wilk normality test. All data were normally distributed (P > .05), and the statistical analysis was performed. A general linear model for repeated measure was applied to assess significant effect of the experimental conditions (road transport, exercise, and acupuncture treatment) on studied parameters. When significant differences were found, Bonferroni’s post hoc comparison was applied. P values <.05 were considered statistically significant. Statistical analysis was performed using the Statistica software package (Statistica 7, Stat Software Inc, Tulsa, OK). 3. Results Statistical analysis revealed a significant effect of transport and exercise (P < .01) on all studied parameters, except for Ca and TSKIN of neck (P > .05), in CG and AG. Higher Hct and TP, and lower Mg, P, and Cl values were
found after transport and exercise respect to rest conditions (Fig. 1). The values of TRECTAL and TSKIN concerning shoulder, ribs, flank, internal thigh, and back regions were higher after transport and exercise respect to rest conditions (Fig. 2). No significant effect of acupuncture stimulation was found on hematochemical parameters (P > .05), whereas it influenced (P < .01) the values of TRECTAL and of TSKIN of flank. As showed in Fig. 2, higher values of TRECTAL and flank TSKIN were found in transported and exercised horses subjected to acupuncture stimulation respect to control horses. 4. Discussion Under stressful stimuli as road transport and physical exercise, the body must find a new dynamic equilibrium and this requires several adaptive body responses. The results obtained in this study showed a significant increase in TP and Hct values immediately after transport and exercise in Thoroughbred horses suggesting a dehydration status of transported and exercised horses. The rise of TP and Hct values following physical effort is explained by the loss of extracellular fluid, decrease of blood volume, hemoconcentration, catecholamine release, and splenic contraction [10,22,23]. The TP increase together with the rise in Hct values is indicative of the fluid shift from the vascular compartment to the interstitial space in order to provide water for sweat production. The sweat loss causes a net reduction in total body water and a decrease of plasma volume [1]. This dynamic exchange of fluid between compartments inevitably leads to a movement of electrolytes allowing the horse to defend the internal environment of the cells. In the present study, a mild decrease in Mg, P, and Cl values was observed following transport and exercise that could be caused by their intracellular movements, which is necessary for their use in muscle function, rather than from their loss with the sweating [1]. It is known that all the electrolytes contribute to the osmotic concentration of the body fluids, a variable that is tightly regulated to prevent cell dehydration or cell swelling [1]. Under
M. Rizzo et al. / Journal of Equine Veterinary Science 55 (2017) 84–89
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Fig. 1. Mean values standard deviation (SD) of hematocrit (Hct), total proteins (TPs), magnesium (Mg), calcium (Ca), phosphorus (P), and chloride (Cl) obtained from acupunctured group (AG) and control group (CG) during transport (at rest: TPRE; after unloading from the vehicle: TPOST; 30 minutes after unloaded: TPOST30) and exercise (at rest: RPRE; at the end of the race: RPOST; 30 minutes after the race: RPOST30).
stressful conditions as transport and exercise, early fluid and electrolyte shifts appear more related to a system-wide redistribution of blood and fluid from capacitance vessels and the interstitial space in order to increase venous return and augment cardiac output. Later responses provide fluid and electrolytes for the production of sweat and thermoregulation. The dynamic flux of fluid provides for removal of the metabolic waste products and for removal of heat produced by working muscles during physical effort [1]. The results obtained in this study showed a significant increase of TECTAL after transport and physical exercise in both AG and CG. During transport, horses’ muscles work in order to cope with changing forces due, for example, to acceleration, to vehicle movements during cornering, and to deceleration [24,25]. During exercise, horses’ muscles work to gallop and a conversion of stored chemical energy into mechanical energy and thermal energy occurs [1,26]. The heat produced by muscle during transport and exercise is such as to raise core temperature [27]. Therefore, heat dissipation mechanisms should be activated. The primary
physiologic mechanism leading to heat dissipation is represented by the increase in cardiac output that brings to heat transfer from core to the cutaneous blood circulation [28]. During transport and exercise, the muscle activity created more internal body heat, which elicited reflex neurogenic vasodilatation in the skin resulting in increased skin temperature [27]. This agrees with the increase of all considered TSKIN found during transport and exercise in both AG and CG with the exception of TSKIN values of the neck region that not showed variations throughout the monitoring period. No influence of acupuncture was found on Hct, TP, and electrolytes values in transported and exercised horses, whereas statistically significant higher values of TRECTAL and TSKIN of flank region were found in horses subjected to acupuncture treatment. Moreover, although not statistically significant, the TSKIN temperature of the other considered regions was higher in acupunctured horses respect to control. The mechanism of the thermoregulatory action of acupuncture stimulation has to be fully
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M. Rizzo et al. / Journal of Equine Veterinary Science 55 (2017) 84–89
Fig. 2. Mean values standard deviation (SD) of rectal (TRECTAL) and skin (TSKIN) temperatures measured in acupunctured group (AG) and control group (CG) during transport (at rest: TPRE; after unloading from the vehicle: TPOST; 30 minutes after unloaded: TPOST30) and exercise (at rest: RPRE; at the end of the race: RPOST; 30 minutes after the race: RPOST30).
understood; however, an increase of body and surface temperature by acupuncture treatment has been reported by many authors [29–31]. It has been stated that acupuncture has an adjusting and controlling effect on some body functions, including the heart rate and blood flow, allowing the physiological activity of the body to maintain homeostasis [32]. Acupuncture treatment could regulate autonomic nerve activities by decreasing the activities of sympathetic function and manifested as vasodilatation, blood flow increase, and, consequently, skin temperature increase [33]. Effectively, it has been established that acupuncture can not only affect the general circulation [34,35], but also affect skin microcirculation [36]. Studies carried out on athlete humans reported that rising body temperature could not only increase the enzyme activities, muscular blood volume, and oxygen intake level but also increase the time between contraction and relaxation of muscles, thus enhancing physical performance and avoiding injuries [37,38]. 5. Conclusion This study demonstrated that Thoroughbred horses exhibited a classic hemodynamic response to stressors as road transport and exercise and that acupuncture treatment affects body temperature and skin temperature of flank region in both transported and exercised horses. A solid scientific framework is missing to analysis the precise mechanisms by which acupuncture affects
thermoregulatory mechanisms. In this study, horses subjected to acupuncture treatment showed a rise in body temperature suggesting an adjusting and controlling effect of acupuncture on thermoregulatory function, probably, by decreasing the activities of sympathetic function and manifested as vasodilatation, arterial blood flow, and skin microcirculation increase. However, other acupuncture points and types of stimulation should be tested to better verify the beneficial effect of this therapy to reduce stress in horses. Future studies in these issues are necessary to investigate the underlying mechanism and its clinical relevance.
References [1] Hinchcliff KW, Kaneps AJ, Geor RJ. Equine sports medicine and surgery. London, UK: Saunders; 2004. [2] Padalino B. Effects of the different transport phases on equine health status, behavior, and welfare: a review. J Vet Behav 2015;10:272–82. [3] Fazio F, Assenza A, Tosto F, Casella S, Piccione G, Caola G. Modifications of some acute phase proteins and the white blood cell count in Thoroughbreds during training. Vet Rec 2010;167:370–2. [4] Padalino B. Effect of transport and exercise on behavior of sport horses. In: Paz-Silva A, Arias Vázquez MS, Sánchez-Andrade Fernández R, editors. Horses: breeding, health disorders and effects on performance and behavior. New York, USA: Nova Science Publishers Inc; 2014. p. 21–44. [5] Turner TA. Diagnostic thermography. Vet Clin North Am Equine Pract 2001;17:95–113. [6] Brody S. Bioenergetics and growth. New York: Reinhold Pub Corp; 1945. p. 902. [7] Li HX. Physiology. Zhengzhou, China: Henan Medical University Publishing House; 1997.
M. Rizzo et al. / Journal of Equine Veterinary Science 55 (2017) 84–89 [8] Charkoudian N. Mechanisms and modifiers of reflex induced cutaneous vasodilatation and vasoconstriction in humans. J Appl Physiol 2010;109:1221–8. [9] Assenza A, Bergero D, Congiu F, Tosto F, Giannetto C, Piccione G. Evaluation of serum electrolytes and blood lactate concentration during repeated maximal exercise in horse. J Equine Vet Sci 2014; 34:1175–80. [10] Piccione G, Giannetto C, Assenza A, Fazio F, Caola G. Serum electrolyte and protein modification during different workload in jumper horse. Comp Clin Pathol 2007;16:103–7. [11] Barlas P, Robinson J, Allen J, Baxter GD. Lack of effect of acupuncture upon signs and symptoms of delayed onset muscle soreness. Clin Physiol 2000;20:449–56. [12] Knardahl S, Elam M, Olausson B, Wallin BG. Sympathetic nerve activity after acupuncture in humans. Pain 1998;75:19–25. [13] Sandberg MT, Lundeberg T, Lindberg LG, Gerdle B. Effects of acupuncture on skin and muscle blood flow in healthy subjects. Eur J Appl Physiol 2003;90:114–9. [14] Tsuchiya M, Sato EF, Inoue M, Asada A. Acupuncture enhances generation of nitric oxide and increases local circulation. Anesth Analg 2007;104:301–7. [15] Jou NT, Ma SX. Responses of nitric oxide-cGMP release in acupuncture point to electroacupuncture in human skin in vivo using dermal microdialysis. Microcirculation 2009;16:434–43. [16] Shinbara H, Okubo M, Kimura K, Mizunuma K, Sumiya E. Participation of calcitonin gene related peptide released via axon reflex in the local increase in muscle blood flow following manual acupuncture. Acupunct Med 2013;31:81–7. [17] Knowles RG, Moncada S. Nitric oxide as a signal in blood vessels. Trends Biochem Sci 1992;17:399–402. [18] Min S, Lee H, Kim S, Park J, Chae Y, Lee H, Park H. Local changes in microcirculation and the analgesic effects of acupuncture: a laser Doppler perfusion imaging study. J Altern Complement Med 2015; 21:46–52. [19] Li W, Ahn A. Effect of acupuncture manipulations at LI4 or LI11 on blood flow and skin temperature. J Acupunct Meridian Stud 2016;9:128–33. [20] Hana YJ, Yi SY, Lee YJ, Kim KH, Kim EJ, Lee SD. Quantification of the parameters of twisting-rotating acupuncture manipulation using a needle force measurement system. Integr Med Res 2015;4:57–65. [21] Lee SY, Son YN, Choi IH, Shin KM, Kim KS, Lee SD. Quantitative study of acupuncture manipulation of lifting-thrusting using a needle insertion-measurement system in phantom tissue. J Korean Med 2014;35:74–82. [22] Jablonska EM, Ziolkowska SM, Gill J, Szykula R, Faff J. Changes in some haematological and metabolic indices in young horses during the first year of jump-training. Equine Vet J 1991; 23:309–11. [23] Kurosawa M, Nagata S, Takeda F, Mima K, Hiraga A, Kai M, Taya K. Plasma catecholamine, adrenocorticotropin and cortisol responses
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Contents lists available at ScienceDirect
Journal of Equine Veterinary Science journal homepage: www.j-evs.com
Original Research
Core and Surface Temperature Modification During Road Transport and Physical Exercise in Horse After Acupuncture Needle Stimulation Maria Rizzo a, Francesca Arfuso a, Elisabetta Giudice b, Francesco Abbate a, Francesco Longo c, Giuseppe Piccione a, * a
Department of Veterinary Sciences, University of Messina, Polo Universitario dell’Annunziata, Messina, Italy Department of Chemical, Biological, Pharmaceutical and Environmental Sciences, University of Messina, Messina, Italy c Veterinary Practitioner, Bologna, Italy b
a r t i c l e i n f o
a b s t r a c t
Article history: Received 7 February 2017 Received in revised form 30 March 2017 Accepted 30 March 2017 Available online 18 April 2017
In this study, the effect of acupuncture on skin temperature (TSKIN), including six body regions (neck, shoulder, ribs, flank, internal thigh, and back), rectal temperature (TRECTAL), serum levels of total proteins, hematocrit, magnesium, calcium (Ca), phosphorus, and chloride was evaluated in five Thoroughbred horses. Horses competed in two official races. For each race, animals were transported from their stables to the racetrack. Horses transported and competed in the first race represent the control group. Two weeks later, the same horses competed in the second race. Before road transport, they were treated with acupuncture. From animals, the TSKIN, TRECTAL, and blood samples were collected at rest; after unloaded, 30 minutes after unloaded, at rest in the transit box, at the end of the race, and 30 minutes after the race. A significant effect of transport and exercise was found on all studied parameters (P < .01), except for Ca and TSKIN of neck (P > .05), in both groups. Transported and exercised horses subjected to acupuncture treatment showed statistically significant higher values of TRECTAL and TSKIN of flank (P < .01). Thoroughbred horses display a classic hemodynamic response to transport and exercise; acupuncture affects body temperature and skin temperature of flank region in both transported and exercised horses suggesting a controlling effect of acupuncture on thermoregulatory function, probably, by decreasing the activities of sympathetic function and manifested as vasodilatation, arterial blood flow, and skin microcirculation increase. Ó 2017 Elsevier Inc. All rights reserved.
Keywords: Acupuncture Body temperature Exercise Horse Road transport
1. Introduction Transport and physical exercise represent stressful stimuli which can lead to homeostasis disruption with direct impact on animal health status and physical
Conflict of interest: The authors disclaim any financial support or relationships that may pose conflict of interest. * Corresponding author at: Giuseppe Piccione, Department of Veterinary Sciences, University of Messina, Polo Universitario dell’Annunziata, 98168, Messina, Italy. E-mail address: [email protected] (G. Piccione). 0737-0806/$ – see front matter Ó 2017 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.jevs.2017.03.224
performance of the athletic horse [1–3]. Athlete horses are used to travel before competitions, but, after transport, some of them show lower performance than usual [4]. To limit health problems related to transport and exercise stress, it is important to evaluate the health status of the horses before and after the physical effort, to optimize the environmental conditions inside the truck or to provide them with electrolytes and antioxidants. The evaluation of body temperature represents a valuable tool to monitor the physiologic status, welfare, and the stress response of animals. Monitoring horse surface temperature allows the evaluation of function of individual parts of the body. The
M. Rizzo et al. / Journal of Equine Veterinary Science 55 (2017) 84–89
assessment of horse surface temperature changes resulting from transport and/or exercise could allow detecting inflamed areas caused by stress of musculoskeletal system that could account for a decreased welfare and physical performance [5]. During exercise, about 20% of the metabolism in the muscle cells is used for work and the remaining 80% becomes heat [6]. Similarity to human, skin is the only heat dissipation way of the equine body; when the body energy metabolism and heat production increases, heat dissipation through the skin increases, too, and so the surface temperature rises [7]. Heat removal rate is generally determined by the heat transfer rate from the core to the surface of the body through blood vessels. In particular, vasodilation allows more blood to flow through the arteriole, thus venting the heat by convection and conduction through the superficial capillaries, if vasodilatation is not sufficient sweating starts [8]. Sweating can lead to fluid and electrolyte losses that if uncompensated, can lead to thermoregulatory instability, hypovolemia, imbalance in electrolyte serum concentration, and when the deficit becomes serious, horses can show clinical signs of dehydration compromising the physical performance [1,9]. Sweat loss of electrolyte, in particular, causes the onset of peripheral fatigue and weakness [10]. It has been demonstrated that acupuncture treatment leads to alleviation of muscle tension, improvement of local blood flow, increase of pain threshold, and modulation of autonomic nervous system [11,12]. Acupuncture stimulation induces alterations in microcirculation, including the increase in the skin and muscle blood flow [13], and it lead to the release of biological factors, such as nitric oxide [14,15] and calcitonin gene-related factor [16], which play important roles in the modulation of vasodilation [17]. It has been well established that acupuncture can influence skin temperature and blood flow in human [18,19]. However, to the best authors’ knowledge, no studies have been currently published on the influence of acupuncture treatment on thermoregulation and hydration status of equine species. Therefore, the aim of the present study was to evaluate how the values of rectal and skin temperatures as well as hydration indices change following acupuncture stimulation in Thoroughbred horses before and after road transport and physical exercise. 2. Materials and Methods 2.1. Animals and Study Design All treatments, housing and animal care reported below were carried out in accordance with the standards recommended by the EU Directive 2010/63/EU for animal experiments. Five clinically healthy and regularly trained Thoroughbred horses (five geldings, mean age 4 1 year; mean body weight 437 15 kg) were used with the informed consent of the owners. Animals were stabled in individual boxes (3.5 3.5 m) at the same training center located in Sicily, Italy (38 000 4900 N, 15 2501800 E, 80 m above sea level) under natural photoperiod (sunrise at 6:11 AM, sunset at 5:13 PM; mean temperature 23 C, relative humidity 70%). Horses were fed, twice a day (7 AM and 5 PM),
85
a total food amount of about 2.5% of horse body weight (forage-to-concentrate ratio 70:30), and water was available ad libitum. The horses competed in two official 1,300meter races at “Mediterraneo” racetrack (Siracusa, Sicily, Italy). For each race, the five horses were transported from their stables to the racetrack in a 5-horse truck. Each animal traveled, tethered with a 50-cm rope on each side of the halter, in an individual tie box (length 2.3 m; width 0.85 m), giving a total space of about 2 m2 and made two journeys of 145 km. Each journey was made 2 weeks after the previous one. The driver and the route were always the same. Thermal and hygrometric records were carried out inside the box for the whole study by means of a data logger (Gemini, UK). The same instrument was placed approximately 5 cm from the center of the ceiling in the van during transport in order to measure the ambient thermal and hygrometric records inside the truck throughout the transport. The temperature in the box ranged between 24 C and 16 C (maximum/minimum), whereas the relative humidity ranged between 25 C and 67%. Temperatures inside the truck ranged between 24 C and 21.5 C (maximum/minimum). The relative humidity inside the truck ranged between 68% and 75%. Horses transported and competed in the first official race represent the control group (CG). Two weeks later, the same horses competed in the second official race. Before road transport, they were subjected to acupuncture treatment (acupuncture group, AG) and were loaded after 30 minutes onto the truck calmly and without the use of force. Acupuncture was performed once a day for 30 minutes across all animals. A stainless-steel needle (diameter 0.25 mm, length 30 mm, Dongbang Acupuncture Inc, Qingdao, China) was inserted vertically into a depth of 2–3 cm, and stimulation was produced by bidirectional twisting of needles, as described in the previous studies [20,21]. In particular, Table 1 shows the name and location of the selected acupoints received from acupuncture group (AG). For acupuncture stimulation, stainless needles were inserted into the left or right side of selected acupoints. The locations of acupoints were found according to the anatomical structures. 2.2. Body Temperature Measurement From each animal, rectal and skin temperatures were measured by the same operator at rest in their box at 8:30 AM; after being unloaded from the vehicle and housed in the transit boxes, 30 minutes after unloaded, at rest in the transit box, at the end of the race, and 30 minutes after the race. Rectal temperature (TRECTAL), taken as representative of body temperature, was measured by a means of a digital thermometer (HI92704, Hanna Instruments Bedfordshire, UK), inserted 15 cm in the rectum. Skin temperature (TSKIN) was measured at six locations (neck, shoulder, ribs, flank, internal thigh, and back) with an infrared thermometer (Testo 826-T1) having a sensitivity of 0.2 C. 2.3. Blood Sampling and Laboratory Analysis From each animal, blood samples were collected by the same operator at the same time points of the body
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Table 1 Name and location of the acupoints selected for the study. Chinese Name
Other Name Location
Ming-men (life gate)
GV 4
Da-zhui (big vertebra)
GV 14
Dan-zhong (chest center)
CV 17
Qian-chan-wan (thoracic fetlock)
SI 3
Shen-mai (extending vessel) Shang-liao (upper bone hole) Ci-liao (second bone hole) Zhong-liao (central bone hole) Xia-liao (lower bone hole) Huang-men (vitals gate) Fei-pan (lung hugging) Hua Tuo Jia Ji (Hua Tuo’s paravertebral points)
BL BL BL BL BL BL
a
62 31 32 33 34 52
In the depression along the dorsal midline at the intervertebral space between L2 and L3 In the depression along the dorsal midline at the cervicothoracic intervertebral space (C7–T1) On the ventral midline, at the level of the caudal border of the elbow, or at the level of fourth intercostal space In a depression on the caudolateral border of the cannon bone (third metacarpal), distal to the end of the lateral splint bone (fourth metacarpal), proximal to the fetlock, over the lateral palmar digital vein In a depression distal to the lateral malleolus First sacral intervertebral space (S1–S2), 1.5 cuna lateral to the dorsal midline Second sacral intervertebral space (S2–S3), 1.5 cuna lateral to the dorsal midline Third sacral intervertebral space (S3–S4), 1.5 cuna lateral to the dorsal midline Fourth sacral intervertebral space (S4–S5), 1.5 cuna lateral to the dorsal midline At the level of the lower border of the spinous process of L1, 3 cuna lateral to the GV-5. On the caudal border of the scapula, 1/3 of the distance from dorsal to the ventral 0.5–1 cuna lateral to the depressions below the spinous processes of the L2–L6.
The cun is a relative or proportional measurement unit, which is used to locate the Jing-Luo and acupuncture points.
temperature recording. Blood samples were collected by jugular venipuncture into 2-mL vacutainer tubes containing ethylenediaminetetraacetic acid and into 8-mL vacutainer tubes with cloth activator (Terumo Co, Tokyo, Japan). Immediately after collection, blood samples were placed in refrigerated bags and transported to the laboratory for the analysis. ethylenediaminetetraacetic acid whole blood samples were processed in the laboratory within 2 hours for the evaluation of hematocrit (Hct) by means of an automated hematology analyzer (HeCo Vet C, SEAC, Florence, Italy). Blood samples collected into tubes containing clot activator were centrifuged at 1,300g for 10 minutes, and the obtained sera were analyzed within 2 hours to estimate the concentration of total proteins (TPs), magnesium (Mg), calcium (Ca), phosphorus (P), and chloride (Cl) using commercially available kits by means of an automated analyzer UV Spectrophotometer (model Slim SEAC, Florence, Italy). 2.4. Statistical Analysis Data, expressed as mean values standard deviation, were tested for normality using the Shapiro–Wilk normality test. All data were normally distributed (P > .05), and the statistical analysis was performed. A general linear model for repeated measure was applied to assess significant effect of the experimental conditions (road transport, exercise, and acupuncture treatment) on studied parameters. When significant differences were found, Bonferroni’s post hoc comparison was applied. P values <.05 were considered statistically significant. Statistical analysis was performed using the Statistica software package (Statistica 7, Stat Software Inc, Tulsa, OK). 3. Results Statistical analysis revealed a significant effect of transport and exercise (P < .01) on all studied parameters, except for Ca and TSKIN of neck (P > .05), in CG and AG. Higher Hct and TP, and lower Mg, P, and Cl values were
found after transport and exercise respect to rest conditions (Fig. 1). The values of TRECTAL and TSKIN concerning shoulder, ribs, flank, internal thigh, and back regions were higher after transport and exercise respect to rest conditions (Fig. 2). No significant effect of acupuncture stimulation was found on hematochemical parameters (P > .05), whereas it influenced (P < .01) the values of TRECTAL and of TSKIN of flank. As showed in Fig. 2, higher values of TRECTAL and flank TSKIN were found in transported and exercised horses subjected to acupuncture stimulation respect to control horses. 4. Discussion Under stressful stimuli as road transport and physical exercise, the body must find a new dynamic equilibrium and this requires several adaptive body responses. The results obtained in this study showed a significant increase in TP and Hct values immediately after transport and exercise in Thoroughbred horses suggesting a dehydration status of transported and exercised horses. The rise of TP and Hct values following physical effort is explained by the loss of extracellular fluid, decrease of blood volume, hemoconcentration, catecholamine release, and splenic contraction [10,22,23]. The TP increase together with the rise in Hct values is indicative of the fluid shift from the vascular compartment to the interstitial space in order to provide water for sweat production. The sweat loss causes a net reduction in total body water and a decrease of plasma volume [1]. This dynamic exchange of fluid between compartments inevitably leads to a movement of electrolytes allowing the horse to defend the internal environment of the cells. In the present study, a mild decrease in Mg, P, and Cl values was observed following transport and exercise that could be caused by their intracellular movements, which is necessary for their use in muscle function, rather than from their loss with the sweating [1]. It is known that all the electrolytes contribute to the osmotic concentration of the body fluids, a variable that is tightly regulated to prevent cell dehydration or cell swelling [1]. Under
M. Rizzo et al. / Journal of Equine Veterinary Science 55 (2017) 84–89
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Fig. 1. Mean values standard deviation (SD) of hematocrit (Hct), total proteins (TPs), magnesium (Mg), calcium (Ca), phosphorus (P), and chloride (Cl) obtained from acupunctured group (AG) and control group (CG) during transport (at rest: TPRE; after unloading from the vehicle: TPOST; 30 minutes after unloaded: TPOST30) and exercise (at rest: RPRE; at the end of the race: RPOST; 30 minutes after the race: RPOST30).
stressful conditions as transport and exercise, early fluid and electrolyte shifts appear more related to a system-wide redistribution of blood and fluid from capacitance vessels and the interstitial space in order to increase venous return and augment cardiac output. Later responses provide fluid and electrolytes for the production of sweat and thermoregulation. The dynamic flux of fluid provides for removal of the metabolic waste products and for removal of heat produced by working muscles during physical effort [1]. The results obtained in this study showed a significant increase of TECTAL after transport and physical exercise in both AG and CG. During transport, horses’ muscles work in order to cope with changing forces due, for example, to acceleration, to vehicle movements during cornering, and to deceleration [24,25]. During exercise, horses’ muscles work to gallop and a conversion of stored chemical energy into mechanical energy and thermal energy occurs [1,26]. The heat produced by muscle during transport and exercise is such as to raise core temperature [27]. Therefore, heat dissipation mechanisms should be activated. The primary
physiologic mechanism leading to heat dissipation is represented by the increase in cardiac output that brings to heat transfer from core to the cutaneous blood circulation [28]. During transport and exercise, the muscle activity created more internal body heat, which elicited reflex neurogenic vasodilatation in the skin resulting in increased skin temperature [27]. This agrees with the increase of all considered TSKIN found during transport and exercise in both AG and CG with the exception of TSKIN values of the neck region that not showed variations throughout the monitoring period. No influence of acupuncture was found on Hct, TP, and electrolytes values in transported and exercised horses, whereas statistically significant higher values of TRECTAL and TSKIN of flank region were found in horses subjected to acupuncture treatment. Moreover, although not statistically significant, the TSKIN temperature of the other considered regions was higher in acupunctured horses respect to control. The mechanism of the thermoregulatory action of acupuncture stimulation has to be fully
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Fig. 2. Mean values standard deviation (SD) of rectal (TRECTAL) and skin (TSKIN) temperatures measured in acupunctured group (AG) and control group (CG) during transport (at rest: TPRE; after unloading from the vehicle: TPOST; 30 minutes after unloaded: TPOST30) and exercise (at rest: RPRE; at the end of the race: RPOST; 30 minutes after the race: RPOST30).
understood; however, an increase of body and surface temperature by acupuncture treatment has been reported by many authors [29–31]. It has been stated that acupuncture has an adjusting and controlling effect on some body functions, including the heart rate and blood flow, allowing the physiological activity of the body to maintain homeostasis [32]. Acupuncture treatment could regulate autonomic nerve activities by decreasing the activities of sympathetic function and manifested as vasodilatation, blood flow increase, and, consequently, skin temperature increase [33]. Effectively, it has been established that acupuncture can not only affect the general circulation [34,35], but also affect skin microcirculation [36]. Studies carried out on athlete humans reported that rising body temperature could not only increase the enzyme activities, muscular blood volume, and oxygen intake level but also increase the time between contraction and relaxation of muscles, thus enhancing physical performance and avoiding injuries [37,38]. 5. Conclusion This study demonstrated that Thoroughbred horses exhibited a classic hemodynamic response to stressors as road transport and exercise and that acupuncture treatment affects body temperature and skin temperature of flank region in both transported and exercised horses. A solid scientific framework is missing to analysis the precise mechanisms by which acupuncture affects
thermoregulatory mechanisms. In this study, horses subjected to acupuncture treatment showed a rise in body temperature suggesting an adjusting and controlling effect of acupuncture on thermoregulatory function, probably, by decreasing the activities of sympathetic function and manifested as vasodilatation, arterial blood flow, and skin microcirculation increase. However, other acupuncture points and types of stimulation should be tested to better verify the beneficial effect of this therapy to reduce stress in horses. Future studies in these issues are necessary to investigate the underlying mechanism and its clinical relevance.
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