Effect of dexamethasone administration on cortisol concentration and biochemical profile in buffaloes suffering from dystocia

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1 Anim. Reprod., v.2, n.4, p , Oct./Dec Effect of dexmethsone dministrtion on cortisol concentrtion nd iochemicl profile in uffloes suffering from dystoci A. Sthy 1, S. Prhkr, S.P.S. Ghumn Deprtment of Animl Reproduction, Gynecology nd Ostetrics, Punj Agriculturl University, Ludhin, , Indi. Astrct The present study ws undertken to evlute drenl response nd iochemicl chnges in the lood following dexmethsone (DEX) dministrtion t the time of prturition nd during the immedite postprtum period in uffloes suffering from dystoci. Plsm cortisol concentrtion, lood glucose, plsm totl proteins, nd non-esterified ftty cids (NEFA) were evluted. Plsm cortisol concentrtion ws higher (P < 0.01) in dystoci groups thn the eutoci group on the dy of prturition. There ws significnt reduction (P < ) in plsm cortisol concentrtion on Dy 1 postprtum in the DEX-treted group. Dexmethsone tretment mintined higher levels of lood glucose (P < 0.05) nd NEFA (P < to P < 0.05) nd mrginlly lowered the totl protein level. This study demonstrted tht the hypothlmo-pituitry-drenl xis ws responsive to DEX suppression during the immedite postprtum period following dystoci, nd DEX tretment ws found to e eneficil in promoting metolism to meet energy demnds during stress. Keywords: drenl response, uffloes, cortisol, dexmethsone, dystoci. Introduction The process of prturition, though physiologicl, is stressful event (Guidry et l., 1976). Anorml prturition further dds to the stress of clving (Nko nd Grunert, 1990, Prhkr et l., 1999). Endogenous glucocorticoids help in the survivl of nimls following stress. In ddition, supplementtion of exogenous glucocorticoids is recommended to support the drenl response to prevent drenl exhustion due to over stimultion during stressful period (Ferguson nd Hoenig, 2001). Activity of the hypothlmo-pituitry-drenl (HPA) xis is regulted y the levels of glucocorticoid secretion tht vries with the presence or sence of stressors (Dllmn et l., 197). Pregnncy in ewes nd the postprtum period in ewes nd cttle were found to e stress-hyporesponsive periods (Przekop et l., 195; Nnd et l., 199; Smrt et l., 1994; Altemus et l., 1995; Windle et l., 1997; Lightmn et l., 2001). Enhnced glucocorticoid negtive feedck (Schmidt et l., 2005) nd elevted progesterone during pregnncy (Keller-Wood 1 Corresponding uthor: sthyvet@yhoo.com Phone: , Fx: Received: Jnury 5, 2006 Accepted: April 3, 2006 et l., 196) were implicted s meditors in the hyporesponsiveness of the HPA xis. On the contrry, reduced forwrd drive rther thn enhnced negtive feedck (Johnstone et l., 2000) nd reduced vsopressin relese from the prventriculr nucleus (Dougls et l., 2003; M et l., 2005) were lso found to medite the hyporesponsiveness. However, Smith et l. (197) nd Owens et l. (197) oserved reduced dexmethsone (DEX) suppressiility of plsm cortisol nd drenocorticotropic hormone in lte pregnnt nd erly postprtum periods in women, respectively. Stressors mintin the HPA xis in n ctivted stte y ttenuting glucocorticoid negtive feedck until the stressful condition susides (As et l., 2004). Hence, the effectiveness of glucocorticoid feedck vries with the physiologicl stge of reproduction nd presence of stressors. The drenl response to glucocorticoid negtive feedck hs not een reported t the time of prturition ssocited with higher degree of physicl, inflmmtory, or immunologicl stress (i.e. dystoci). Moreover, the persistence of elevted plsm cortisol concentrtions for longer periods in uffloes suffering from dystoci (Prhkr et l., 1999) questioned the effectiveness of negtive feedck ssocited with difficult prturition. The present study ws conducted to evlute the negtive feedck effects of DEX in uffloes suffering from dystoci. In ddition, the iochemicl ltertions following DEX tretment in uffloes suffering from dystoci were lso studied. Mterils nd Methods Animls nd dexmethsone dministrtion Twenty-one uffloes, divided into three groups, were included in the present study. The normlly clving or eutoci group () included 6 normlly-clving Murrh uffloes from the Punj Agriculturl University (PAU) diry frm. The nimls were housed in individul clving pens -15 dys efore prturition nd were mintined under uniform mngement nd feeding conditions. The dystoci control group (, n=5) nd dystoci tretment group (, n=) included prturient Murrh uffloes rought into the Veterinry Clinics of PAU for tretment of dystoci from vrious prts of the stte of

2 Sthy et l. Dexmethsone dministrtion in uffloes suffering from dystoci. Punj, Indi, within hours of onset of prturition (i.e. dominl strining in cses of dystoci due to torsion of uterus or rupture of llntoin without further progress in delivery in other cses ws tken s the time of onset of prturition). These nimls were given the necessitted ostetricl [included mnipultive delivery (: n=1; : n=2), fetotomy (: n=1, : n=2), uterine detorsion (: n=2; : n=3), or cesren section (: n=1; : n=3)] nd supportive [included equl mounts of the sme ntiiotics, sline, nd ecolics] tretments. In ddition to these tretments, uffloes were dministered DEX (40 mg, i.v.; Dexon, Srhi- Zydus, Indi) efore relieving dystoci (Dy 0) nd susequently once dily for two dys postprtum (Dys +1 nd +2). Buffloes in the group were not given DEX tretment. Blood smpling schedule Blood smples were collected dily from uffloes t 0900 h from Dy 3 preprtum (Dy -3) to Dy 3 postprtum (Dy +3). Buffloes suffering from dystoci were smpled thrice on Dy 0 (efore, during nd immeditely fter ostetricl tretment on the dy of ssisted delivery) nd susequently once dily for three dys. In ll the groups, lood smples (5 ml) were collected in heprinized glss vils (50 IU heprin/vil) y jugulr venipuncture efore giving ny tretment. An liquot of 2 ml of lood ws used for preprtion of protein-free filtrte for lood glucose estimtion nd the rest ws centrifuged t 500 g for 15 min for seprtion of plsm, which ws stored t -20 ºC until ssyed for cortisol, totl proteins, nd non-esterified ftty cids (NEFA). Assy procedures Plsm cortisol concentrtions were determined y rdioimmunossy s descried previously (Prksh nd Mdn, 194). Stndrd dilutions of cortisol from ng/ml were used in ech ssy for plotting the stndrd curve. The ssy sensitivity ws 0.2 ng/ml. The intr- nd interssy coefficients of vrition were 9.2% nd.7%, respectively, nd were determined from qulity controls in the rnge of 0.4 to 2 ng/ml. There ws no cross-rectivity with DEX in the ssy. Blood glucose (mg/dl), totl plsm proteins (g/dl), nd NEFA (mg/l) were estimted y Hden s modifiction of the Folin-Wu method (Goldenerg nd Frnkel, 1970), Biuret method (Wootton, 1964), nd spectrophotometric method (Lowry nd Tinsley, 1976), respectively. Sttisticl nlyses Dt re expressed s the men ± SEM. Twowy ANOVA ws done to test for the effects of group nd time nd their interction. When fixed effects were found to e significnt, differences were tested using Duncn s multiple rnge test. Person s correltion ws clculted etween plsm cortisol nd NEFA nd the significnce ws tested using t-test. Anlyses were performed using the Sttisticl Pckge for Socil Sciences softwre progrmme (SPSS 12.0, Chicgo, IL; 2003), nd P < 0.05 ws considered sttisticlly significnt. Plsm cortisol Results In the eutoci group, the plsm cortisol concentrtion on Dy 0 ws higher (P < 0.01) compred to ll of the pre- nd post-prtum dys of oservtion (Fig. 1A). In oth the dystoci groups on Dy 0, plsm cortisol ws higher (P < 0.01) compred to the eutoci group (Fig. 2A). The effect of time ws significnt (P < 0.001) nd decline in cortisol concentrtion ws oserved in ll the groups during the immedite postprtum period. In the, plsm cortisol concentrtion decresed (P < ) from Dy 0 to Dy +1, while the reduction oserved in the ws not significnt. On Dy 0, plsm cortisol concentrtions were not different etween the nd ut were lower (P < 0.05) on Dys +2 nd +3 in the. The percentge decrese in plsm cortisol concentrtion from Dy 0 (fter ostetricl tretment in nd ) to Dy +3 ws 24%, 41%, nd 77% in the,, nd, respectively. Blood glucose, plsm totl proteins, nd non-esterified ftty cids In the eutoci group, lood glucose levels on Dy 0 were higher (P<0.05) thn those on Dys -2 nd -3 (Fig. 1B). In the, the lood glucose level on Dy 0 fter delivery y ostetricl mnipultion ws higher (P < 0.05) thn tht on Dy 0 in the (Fig. 2B). In the, the lood glucose level decresed (P < 0.05) on susequent dys postprtum to levels lmost similr to those of the. In the, the decrese in lood glucose level ws not significnt, nd higher (P < 0.05) level ws mintined compred to the nd. The plsm totl protein level round clving did not vry significntly in the (Fig. 1C). The levels in the nd were pprently lower on ll the dys of oservtion lthough they did not vry significntly from the (Fig. 2C). The protein level on Dy 0 efore ostetricl tretment ws significntly higher (P < 0.05) in the s compred to the due to rndom selection of nimls. Although the protein level in the decresed from Dy 0 to Dy +1, it ws not sttisticlly significnt. In the, the NEFA level on Dy -3 grdully incresed to rech higher (P < 0.05) level on Dy 0 nd then declined y Dy +3 (P < 0.05). The level on Dy 0 ws higher (P < 0.05) thn ll pre nd post-prtum dys (Fig. 1D). Concentrtion of plsm cortisol nd NEFA were lowly correlted (r = 0.307; P < 0.05). In the, 234 Anim. Reprod., v.2, n.4, p , Oct./Dec. 2005

3 Sthy et l. Dexmethsone dministrtion in uffloes suffering from dystoci. the NEFA level on ll the postprtum dys remined higher (P < 0.05) thn tht in the (Fig. 2D). But in the, the NEFA level ws higher (P < 0.05) only on Dy +3 s compred to tht in the. The level in the on Dy +2 nd +3 ws higher (P < 0.05) thn in the A Plsm cortisol (ng/ml) l B Blood glucose (mg/dl) l C Plsm totl proteins (g/dl) l D NEFA (mg/l) l Dys reltive to prturition Figure. 1. Plsm cortisol (ng/ml; A), lood glucose (mg/dl; B), plsm totl proteins (g/dl; C), nd non-esterified ftty cids (mg/l; NEFA; D) in normlly-clving uffloes (eutoci) from Dys -3 to 3 reltive to prturition. Different letters (, ) indicte significnt differences (Plsm cortisol: P < 0.01; Blood glucose nd NEFA: P < 0.05). Anim. Reprod., v.2, n.4, p , Oct./Dec

4 Sthy et l. Dexmethsone dministrtion in uffloes suffering from dystoci. 0 A Plsm cortisol (ng/ml) l B Blood glucose (mg/dl) l C Plsm totl proteins (g/dl) l D NEFA (mg/l) l Before Ost Rx During Ost Rx After Ost Rx Dy +1 Dy +2 Dy +3 Dy 0 Figure. 2. Plsm cortisol (ng/ml; A), lood glucose (mg/dl; B), plsm totl proteins (g/dl; C), nd non-esterified ftty cids (mg/l; NEFA; D) in normlly-clving uffloes (eutoci) nd uffloes suffering from dystoci (control nd tretment) efore, during, nd fter prturition or ostetricl tretment. Downwrd rrows indicte dexmethsone tretments on Dys 0, +1 nd +2 in dystoci tretment uffloes. Ost Rx refers to ostetricl tretment. : Eutoci group; : Dystoci control group; : Dystoci tretment group. Different letters (, ) indicte significnt differences (P < 0.05). 236 Anim. Reprod., v.2, n.4, p , Oct./Dec. 2005

5 Sthy et l. Dexmethsone dministrtion in uffloes suffering from dystoci. Discussion Clving, though physiologicl process, is stressful for oth the dm nd the fetus even in uncomplicted cses (Hudson et l., 1976). In uffloes, the grdul increse in plsm cortisol concentrtion during the preprtum period tht reched to pek on the dy of clving could e explined y preprtum nxiety nd myometril contrctions ssocited with the stress of prturition (Gillette nd Holm, 1963; Nokes et l., 2001). Furthermore, prturition is n inflmmtory process involving the relese of cytokines nd prostglndins, which stimulte the HPA xis (Hlgunset et l., 1994; Kelley, 1996; Rivest, 2001). Therefore, the increse in plsm cortisol concentrtions during the immedite preprtum period could lso e relted to incresing concentrtions of inflmmtory meditors. In uffloes suffering from dystoci, severe hypercortisolemi oserved on the dy of clving could e relted to prolonged stress, musculr ctivity, nd pin. However, the removl of the fetus leds to decrese in plsm cortisol concentrtion during the postprtum period. This decrese in plsm cortisol concentrtion could e due to the removl of the stress of dystoci nd the negtive feedck effect of plsm cortisol (Miller nd O Collghn, 2002). But during the immedite postprtum period, plsm cortisol concentrtions in dystoci control uffloes remined higher thn those in uffloes. A higher degree of inflmmtory response nd stress-response meditors mintin higher plsm cortisol concentrtion despite the negtive feedck effect (As et l., 2004). In uffloes suffering from dystoci, during the postprtum period, the greter decrese in plsm cortisol concentrtion in the DEX-treted group compred to the untreted group could e ttriuted to the negtive feedck effect of DEX ecuse it is cple of more pronounced suppression of the HPA xis thn endogenous cortisol (Ferguson nd Hoenig, 2001). Hence in this study, DEX tretment constrined the stress-induced HPA ctivity in uffloes suffering from dystoci. Such results were lso otined in lortory nimls, nd delyed negtive feedck mechnism, which occurred min fter DEX tretment, ws proposed s cuse (DeKeyser et l., 2000; Ginserg et l., 2003). However, in the present study, DEX pretretment did not suppress the plsm cortisol concentrtion during nd fter ostetricl mnipultions, which vried in durtion from 30 min to 2 h. This could e due to strong drenl stimultion y the stress of dystoci nd ostetricl mnipultions, thus overcoming the immedite potent negtive feedck effects of DEX. The DEX negtive feedck effect ws pronounced on Dy +1 when the plsm cortisol concentrtion declined. However, it ws not suppressed further on susequent dys despite continution of DEX tretment t 24 h intervls up to Dy +2. Hence, DEX tretment overcme the HPA-stimulting effects of inflmmtory meditors to suppress the HPA response on the immedite postprtum dy, ut it could not suppress the cortisol concentrtion therefter. Persisting higher concentrtions of inflmmtory meditors like PGF 2α, (Nko et l., 1997), rective oxygen metolites s indicted y higher level of lipid peroxidtion products (Sthy et l., 2005), nd endotoxemi s result of uterine infections following dystoci (Singh, 2002), might e responsile for reduced suppressiility of plsm cortisol y DEX in the erly postprtum dys, which re cple of ttenuting glucocorticoid negtive feedck (As et l., 2004). Since effects of DEX tretment were not studied in uffloes, the degree of suppression of plsm cortisol y DEX in uffloes suffering from dystoci could not e compred to them. However, this study demonstrted tht glucocorticoid negtive feedck is effective round prturition in uffloes suffering from dystoci, nd the HPA xis is responsive to DEX tretment round the immedite postprtum dy in these uffloes ut not therefter. Cortisol induces the enzymes involved in gluconeogenesis in heptic tissue nd inhiits utiliztion of glucose in extr-heptic tissues, therey elevting the lood glucose concentrtion (Myes nd Bender, 2003). The lood glucose levels were higher in uffloes suffering from dystoci reflecting higher degree of stress. It ws lso oserved tht lood glucose level incresed following vrious ostetricl procedures. Ostetricl mnipultions hve een found to e stressful in the pst (Prhkr et l., 1999). The elevted lood glucose level ws due to elevted cortisol nd ctecholmines, which incresed following vrious ostetricl procedures s response to stress (Brezile, 197 Nko nd Grunert, 1990). Decresing levels of plsm cortisol fter the removl of the fetus nd thus the stress due to dystoci resulted in the decrese of lood glucose levels during the immedite postprtum period. Since DEX is 30 times more potent thn cortisol in its gluconeogenic effects (Ferguson nd Hoenig, 2001), higher lood glucose level ws mintined in DEX treted uffloes. This might e eneficil during cute phses of stress to meet the urgent needs of reestlishing homeostsis nd fueling the essentil orgns. However, prolonged DEX tretment my ntgonize insulin resulting in poor glucose utiliztion (dietes-like condition) nd enhnced ctolic effects in lymphoid nd connective tissue, muscle, ft, nd skin (Ferguson nd Hoenig, 2001). The mrginlly lower levels of totl proteins in uffloes suffering from dystoci, s compred to uffloes, during the immedite postprtum period were due to decrese in protein synthesis s consequence of higher level of plsm cortisol (Ferguson nd Hoenig, 2001). There ws no ltertion in totl protein level due to DEX tretment. Anim. Reprod., v.2, n.4, p , Oct./Dec

6 Sthy et l. Dexmethsone dministrtion in uffloes suffering from dystoci. When fuel is needed for the ody, tricylglycerols stored in dipose tissue re hydrolysed y hormone-sensitive lipses within the dipocytes to relese NEFA (Nelson nd Cox, 2005). Epinephrine gretly stimultes the tricylglycerol lipse, nd the glucocorticoids hve permissive effects for this enzyme ctivity (Ferguson nd Hoenig, 2001). Hence, NEFA re the indictors of n increse in demnd for energy. Higher levels of NEFA in uffloes were due to ctecholmines nd cortisol relesed t the time of prturition s stress response (Axelrod nd Reisine, 194). Dexmethsone mintined higher levels of NEFA in DEX-treted uffloes suffering from dystoci. Incresed NEFA levels re needed under severely stressful conditions (Ferguson nd Hoenig, 2001). Hence, DEX tretment my e dvntgeous in directing the ft reserves to meet the enhnced fuel demnd of the ody during stress. In summry, the responsiveness of the HPA xis to DEX tretment nd effectiveness of glucocorticoid negtive feedck following difficult prturition in uffloes hve een documented in the present study. Since continued DEX tretment for 3 dys did not suppress the plsm cortisol concentrtion eyond Dy +1 postprtum, it my e scertined tht the DEX suppressiility of plsm cortisol ws reduced in uffloes suffering from dystoci during the postprtum period. Higher lood glucose nd NEFA levels in DEX-treted uffloes suggested tht DEX promoted metolic pthwys to meet the required demnd for energy. Hence DEX dministrtion could e eneficil during stressful episodes. However, the effects of higher doses of DEX recommended during stressful periods on HPA suppression, drenl secretory cpcity, immunity, nd insulin ntgonism re yet to e studied in uffloes during the periprturient period. Acknowledgments We re thnkful to: Prof. H. Doson, University of Liverpool, UK, for the supply of nticortisol ntiodies; Dr. G.V.P.P.S. Rvi Kumr, Assistnt Professor, Deprtment of Animl Genetics nd Breeding, Punj Agriculturl University, Ludhin, Indi, for ssistnce in sttisticl nlyses of the dt; nd Dr. G.S. Dhliwl, Professor nd Hed, Deprtment of Animl Reproduction, Gynecology nd Ostetrics, Punj Agriculturl University, Ludhin, Indi, for hving provided the necessry infrstructure for the reserch. The Junior Reserch Fellowship provided y the Indin Council of Agriculturl Reserch to the first uthor is duly cknowledged. References Altemus M, Deuster PA, Glliven E, Crter CS, Gold PW Suppression of hypothlmicpituitry-drenl xis responses to stress in lctting women. J Clin Endocrinol Met, 0: As K, Iwski Y, Yoshid M, Asc M, Oiso Y, Murohr T, Kshimoto K Attenution y rective oxygen species of glucocorticoid suppression on proopiomelnocortin gene expression in pituitry corticotroph cells. Endocrinology, 145: Axelrod J, Reisine TD Stress hormones: their interction nd regultion. Science, 224: Brezile JE Physiologic sis nd consequences of distress in nimls. J Am Vet Med Assoc, 191: Dllmn M, Akn S, Cscio C, Drlington D, Jcoson L, Levin N Regultion of drenocorticotropic hormone secretion: vritions on theme of B. Rec Prog Horm Res, 43: DeKeyser FG, Leker RR, Weidenfeld J Activtion of the drenocorticl xis y surgicl stress: involvement of centrl epinephrine nd interleukin-1. Neuroimmunomodultion, 7:12-1. Dougls AJ, Brunton PJ, Bosch OJ, Russel JA, Neumnn ID Neuroendocrine responses to stress in mice: hyporesponsiveness in pregnncy nd prturition. Endocrinology, 144: Ferguson DC, Hoenig M Glucocorticoid, minerlocorticoid nd steroid synthesis inhiitors. In: Adms HR. (Ed.). Veterinry phrmcology nd therpeutics. th ed. Iow: Iow Stte University Press. pp Goldenerg S, Frnkel S Crohydrtes. In: Frnkel S, Reitmn S, Sonnenwirth AC. (Eds.). Grdwohl s clinicl lortory methods nd digonosis. 7 th ed. Sint Louis: CV Mosy. pp Gillette DD, Holm L Preprtum to postprtum uterine nd dominl contrctions in cows. Am J Physiol, 204: Ginserg AB, Cmpeu S, Dy HE, Spencer RL Acute glucocorticoid pretretment suppresses stress-induced hypothlmo-pituitry-drenl xis hormone secretion nd expression of corticotropic hormone hnrna ut does not ffect c-fos mrna or Fos protein expression in the prventriculr nucleus of the hypothlmus. J Neuroendocrinol, 15:75-3. Guidry AJ, Ppe MJ, Person RE Effects of prturition nd lcttion on lood nd milk cell concentrtions, corticosteroids nd neutrophil phgocytosis in the cow. Am J Vet Res, 37: Hudson S, Mulford M, Whirrlestone WG, Pyne E Bovine plsm corticoids during prturition. J Diry Sci, 59: Hlgunset J, Johnsen H, Kjollesdl AM, Qvigstd E, Espevik T, Austgulen R Cytokine levels in mniotic fluid nd inflmmtory chnges in the plcent from norml deliveries t term. Eur J Ostet Gynecol Reprod Biol, 56: Johnstone HA, Wigger A, Dougls AJ, Neumnn ID, Lndgrf R, Seckl JR, Russell JA Attenution of hypothlmo-pituitry-drenl xis stress responses in lte pregnncy: chnges in feedforwrd nd feedck 23 Anim. Reprod., v.2, n.4, p , Oct./Dec. 2005

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