Page 87 -

โครงการรวบรวมและจัดทําเอกสารวารสารอิเล็กทรอนิกส์ มหาวิทยาลัยเกษตรศาสตร์

76

hormone (FSH) and a single injection of inhibin, a negative feedback of the FSH. Stress is a one of
limiting factor in assisted reproduction (Lian et al., 2013). Several studies have reported the influence
of stress on reproductive function in females (Michael & Cooke, 1994; Kalantaridou et al., 2004; Lian
et al., 2013). The hypothalamic-pituitary-adrenal (HPA) axis activity in the production of
catecholamine and glucocorticoids, which are stimulate corticotropin-releasing hormone (CRH) from

hypothalamus. CRH inhibits hypothalamic gonadotropin-releasing hormone (GnRH) secretion,
subsequently suppresses reproductive activity. In contrast, CRH stimulate pituitary
adrenocorticotropic hormone (ACTH) secretion and subsequently induce glucocorticoids secretion
from adrenal gland. To detect fecal glucocorticoid metabolites, noninvasive method are commonly
used as indicators of biological neuroendocrine response to stress especially in wild animals (Keay
et al., 2006). Moreover, stress including extreme exertion and struggle affects to muscle damage. The
superovulation methods should be designed to eliminate as much handling stress as possible
(Wenkoff & Bringans, 1991).
To our knowledge, no information is available on the consequences of stress and muscle

damage on superovulation response in Rusa deer. Therefore, the major objective of this study was
to examine the effect of restraining on fecal cortisol concentration, hematology and blood chemistry
following two treatments designed to promote superovulation in Rusa deer.

MATERIALS AND METHODS

Five healthy adult female Rusa deer (age 6-7 years old and body weight of 32-45 kg) were

maintained at Kasetsart University, Kamphaeng Saen Campus, Thailand. Animals were separated into
two groups following superovulation protocols; FSH injection group (Group A: restrain 10 times; n=3)
and Anti/Inhibin injection group (Group B: restrain 7 times; n=2) (Figure 1). Superovulation protocols
th
started from 15 January, 2015 (Day 0) and lasted for 20 days. All animal procedures were approved
by the Deer Co-operative of Thailand, LTD. (DCOT).
Blood samples were collected when handling after the animals are safely restrained during
superovulation program. Health and muscle damage parameters were evaluated by hematology and
th
blood chemistry. Fecal samples of all animals were collected every three days during 5 January to
th
16 February, 2015 (Day -10 to Day 32; n=75). Fresh feces were collected between 08.00 to 09.30
hrs. and immediately stored at -20◦C until processing. Frozen fecal samples were dried in an oven
for approximately 72 hours and extracted with ethanol following procedure described by Brown et
°
al. (1994). Extracted samples containing steroid metabolites were stored at –20C until the analysis.
Fecal concentration of cortisol metabolite (antibody: monoclonal anticortisol R 4866) was
determined by enzyme immunoassay as described by Brown et al. (2004). The intra- and inter-assay
coefficients of variation (CV) were 4.59% and 4.96% (n=8), respectively. Data are expressed as ng/g
dry feces.

Fecal concentration of cortisol metabolite was compared using the Repeated Measured
Analysis of Variance (Repeated ANOVA). Data were presented as mean ± SEM. Hematology and blood
chemistry data were analysed using the Statistical Analysis Systems software package (Version 9.0,
SAS Institute Inc., 1996, NC, USA). The values of p<0.05 were considered statistically significant.

วารสารสัตว์ป่าเมืองไทย ปีที่ 24 พ.ศ. 2560 Journal of Wildlife in Thailand Vol. 24, 2017