THE RELATIONSHIP BETWEEN VAGINAL MUCOUS IMPEDANCE AND SERUM CONCENTRATIONS OF ESTRADIOL AND PROGESTERONE THROUGHOUT THE SHEEP ESTROUS CYCLE

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1 ELSEVIER THE RELATIONSHIP BETWEEN VAGINAL MUCOUS IMPEDANCE AND SERUM CONCENTRATIONS OF ESTRADIOL AND PROGESTERONE THROUGHOUT THE SHEEP ESTROUS CYCLE 1 P.M. Bartlewski, A.P. Beard and N.C. Rawlings Department f Veterinary Physilgical Sciences, Western Cllege f Veterinary Medicine University f Saskatchewan, Saskatn, SK, Canada. Received fr publicatin: July 6, 199 Accepted: Octber ].3, 3_99 ABSTRACT The bjective f this experiment was t assess the relatinship between electrical resistance f the vaginal mucsa and serum cncentratins f estradil (E2) and prgesterne (P4) during the estrus cycle in ewes. Vaginal impedance was recrded daily using a 2-electrde impedmeter in 10 nnprlific Western white-faced and 7 prlific Finn ewes, during the mid-breeding seasn (Octber t December). Transrectal ultrasngraphy f varies was perfrmed nce a day t cnfirm vulatin and mnitr fllicle grwth (fllicles ~ 3 mm in diameter) and develpment f crpra lutea (CL). Jugular bld samples were cllected daily fr radiimmtmassay (RIA) f estradil and prgesterne. In all ewes, a decline in vaginal impedance (t <40 hms) was clsely assciated with the nset f behaviral estrus. In bth breeds f sheep, there was n significant crrelatin between dally serum cncentratins festradil and vaginal impedance thrughut the estrus cycle. Daily serum cncentratins f prgesterne and the E2:P 4 rati were crrelated with vaginal impedance during the perid f lutelysis and fllicular phase in bth breeds (Western white-faced ewes: r = 0.62, P = and r = -0.56, P = ; Finn ewes: r = 0.61, P = and r = -0.45, P = 0.03, respectively) and early in the cycle (Days 0 t 2, Day 0 = day f vulatin) in white-faced ewes (r = 0.6 I, P = and r = -0.36, P = 0 052, respectively) but nt during the remaining prtin f the luteal phase in either breed. In cnclusin, vaginal mucus impedance appears t be primarily cntrlled by prgesterne, but it als changes in respnse t shifts in the E2:P 4 rati when prgesterne cncentratins are lw. Impedmetric characteristics f the vaginal mucsa in cyclic ewes are an indicatr f serum cncentratins f prgesterne and E2:P 4 ratis during the terminal stage f the estrus cycle by Elsevier Science Inc. Key wrds: vaginal impedance, estradil, prgesterne, sheep, estrus cycle Acknwledgments This study was funded by the Alberta Agricultural Research Institute, in the frm f a grant t Dr. N.C. Rawlings. Pawel M. Bartlewski was supprted by a University f Saskatchewan Graduate Teaching Fellwship The authrs thank Ms. Susan Ck fr her excellent technical ~csistance. rrespndence and reprint requests: Deparmaent f Veterinary Physilgical Sciences, Western Cllege f Veterinary Medicine, University f Saskatchewan, 52 Campus Drive, Saskatn, SK, S7N 5B4, Canada. Therigenlgy 51:13-27, X/99/$-see frnt matter 1999 by Elsevier Science Inc. PII S X(99)

2 14 Therigenlgy INTRODUCTION Ovarian fllicular grwth and luteal tissue develpment are assciated with histlgical (3, 5, 22) and histchemical (17) changes in the mucsal layer f reprductive rgans, including the vagina. These changes, in turn, are fllwed by alteratins in electrical prperties f the mucsal tissues (2). It has been suggested that these alteratins reflect different secretin rates fr estrgens and prgesterne, and hence may be useful in determining the stage f the estrus cycle (11). In ewes, it has been demnstrated that the highest cnductivity/lwest impedance f the vaginal mucus membrane ccurs just befre estrus, and it remains high/lw fr 24 t 4 h (23, 1, 2). Hwever, the relatinship between daily serum cncentratins f varian sterids and impedmetrie prperties f the vaginal mucsa thrughut the sheep estrus cycle has nt been studied. The aim f this study was t see if electrical resistance f the vaginal muesa near the cervix uteri is crrelated with circulating cncentratins f estradii and prgesterne at different stages f the vulatry cycle in the ewe. We attempted t find mathematical relatinships that wuld allw fr the predictin f serum cncentratins f estradil and prgesterne based n vaginal mucus impedance. During the estrus cycle in the ewe, estradil secretin is related t the grwth f fllicles t an vulatry size f 25 mm in diameter (6), and serum cncentratins f prgesterne are related t the ttal vlume f luteal tissue (unpublished bservatins). We hypthesized that the measurement f vaginal impedance culd serve as an indicatr f the fllicular and luteal status in cyclic ewes. T cmpare the effects f the 2 varian sterids n vaginal impedance, we measured daily vaginal impedance and serum cncentratins f estradil and prgesterne during 1 estrus cycle in 2 breeds f sheep differing in secretin f estradil and prgesterne (nnprlifie Western white-faced and prlific Finn ewes). Animals and Experimental Prcedures MATERIALS AND METHODS Ten Western white-faced (apprximately 5 yr f age, average bdy weight 90 :t: 7 kg) and seven Finn (3 t 4 yr f age, average bdy weight 57 ~- 4 kg) clinically healthy ewes were used in the present study during the mid-breeding seasn (Octber t December). The ewes were hused utdrs in sheltered dry-lts and fed dally maintenance ratins f alfalfa pellets while water, hay and cbalt idized salt licks were available ad libitum. Estrus was detected with 3 vasectmized crayn-harnessed rams. Vaginal mucus impedance was recrded daily using a 2-electrde impedmeter a (shaft length 20.5 cm; shaft diameter 2.0 em; electrde uter diameter 2.2 cm). The instrument was designed and validated fr use in sheep (2). Using this impedmeter, a decline in vaginal mucus impedance belw 40 hms was shwn t ccur at the beginning f behaviral estrns (2). Befre each measurement, the shaft f the impedmeter was cleaned, disinfected and dried. The prbe was then inserted int the vagina, and the end f the shaft with the recrding electrdes was placed in clse prximity and psterir t the uterine cervix. Three cnsecutive readings f impedance were Electrnic Oestrus Detectr; Firma Draminski, Olsztyn, Pland.

3 Therigenlgy 15 taken by switching an n/fftggle befre the prbe was remved frm the vagina, and the lwest value was recrded. Dally transrectal ultrasngraphy f varies was perfrmed t detect days f vulatin and mnitr varian fllicle grwth (fllicles ~ 3 mm in diameter) and develpment f CL. Each animal underwent dally ultrasngraphic examinatin f varies fr 1 estrus cycle, cmmencing n the day n which the ewe was first marked by a ram r a decline in vaginal impedance readings belw 40 hms was recrded. Days f vulatin were regarded as the days n which large varian antral fllicles that had been identi~ed by ultrasngraphy were n lnger seen. Ultrasngraphy utilized a B-mde, real-time searmer equipped with a 7.5 MHz haman prstate transducer (shaft length 35 cm; shaft diameter 1.6 cm) validated fr the present applicatin in ewes (19, 25), and was perfrmed by an experienced peratr. Bld samples (10 ml) were cllected daily. by jugular.. venlpuncture.. int vacutalners c and allwed t clt fr 1 t 24 h at rm temperature. After centrifugatin at 3000 rpm (2000 x g) fr 15 min, serum was harvested and stred at -20 C fr analysis at a later date. Hrmne Assays Serum samples were analyzed by RIA fr cncentratins festradil (13) and prgesterne (21). The sensitivities f the assays were defined as the lwest cncentratins f standard that differed frm zer (unpaired t-test, P<0.05). The sensitivities f the assays were 1.0 pg/ml and 0.03 ng/ml fr estradil and prgesterne, respectively. Serum samples with cncentratins f hrmnes belw assay sensitivity were assigned a cncentratin equal t the sensitivity f the assay. The cncentratins f standards ranged frm 1.0 t 100 pg/ml (estradil) and frm 10 pg/ml t 10 ng/ml (prgesterne). Fr reference sera with mean estradil cncentratins f 9.3 r 1.9 pg/ml, the intra- and inter-assay cefficients f variatin (CVs) were 15.0 and 10.1%, r 16.3 and 10.9%, respectively. Intra- and inter-assay CVs fr vine reference sera with mean prgesterne cncentratins f 0.49 r 1.56 ng/ml were 11.9 and 4.2%, r 14. and 11.2%, respectively. Statistical Analysis Dally values fr vaginal mucus impedance, serum cncentratins f estradil and prgesterne, and the E2:P 4 rati were nrmalized t the day f the first vulatin (Day 0) fr the perid frm Day - 1 t Day 17 f the intervulatry interval studied, and were analyzed fr day and breed effects and day-by-breed in, actins using multivariate analysis f variance (General Linear Mdel prcedures in MINITAB--). If the main effect f day was significant, Fisher's prtected least significant difference (LSD) was used as the pst-anova test t cmpare individual (daily) means within each breed. If there was a significant effect f breed r day-by-breed interactin, differences between breeds within each day were determined by a t-test. Daily serum cncentratins b Alka SSD 500 Ech Camera, Overseas Mnitr Crp. Ltd., Richmnd, BC, Canada. d Bectn Dica~sn, Rutherfrd, N J, USA. MINITAB v, Statistical Sftware, Minitab Inc., Enhanced Versin, Release 9.1 fr VMS/VAX 6.2; 1992, State Cllege, PA, USA.

4 16 Therigenlgy El 70 (a) 70 (b) 6O O 20 ' I ' I ~ ' I ' I ' I ' I ' t ' I ' ] 20 ' I ' I ' I ' I ' I ' I ' I ' I ' I ' I IO G e~ v s ~ u 2 2 N -2 0 i I I I I I I ' ~ ~ ! ~ q ~-I ' I ' q ' I ' I ' I ' I ' i ' I ~1 ' I ' I 7 I r V' I ' I ~ Days frm first vulatin f Lhe inlervulalry interval Figure 1. Daily vaginal mucus impedance (tp panels, II), serum cncentratins f estradil (A) and prgesterne (-bld, middle panels), and estradil:prgesterne ratis (bttm panels, t), in 2 Western white-faced ewes (a-b) studied during 1 estrus cycle in the middle prtin f the breeding seasn (Octber t December).

5 Therigenlgy (a) 70 (b) ~ ~. 50 II I ~ ~ " I~ ~" "~ 4 ~ 3. l i.d 0 t I ' i r~-i, I ' --~ r "~ s ~ '~, I000 I000 I00 lo0 ~ lo 1.I ' i ' r ~ i ' I ' I,--I, I ' J ' I ' I S S Days frm first vulatin f the intervulatry interval Figure 2. Daily vaginal mucus impedance (tp panels, ra), serum cncentratins festradil (A) and prgesterne (-, middle panels), and estradil:prgesterne ratis (bttm panels, 0), in 2 Finn ewes (a-b) studied during 1 estrus cycle in the middle prtin f the breeding seasn (Octber t December).

6 1 Therigenlgy f prgesterne and estradil, the E2:P 4 rati and vaginal mucus impedance were als aligned t the day f the secnd vulatin f the intervulatry interval (Day 0), and mean values were determined fr the perid frm Day - 5 t Day 0, fr bth breeds f sheep studied. Differences between breeds and between days within each breed were analyzed by analyses f variance, as described abve. All results are given as mean ± SEM. T study the relatinship between vaginal mucus impedance and peripheral cncentratins f varian sterids, the estrus cycle was divided int 4 perids, based n the physical characteristics f CL and changes in mean serum cncentratins f prgesterne. Because estradil secretin during the estrus cycle in individual ewes appeared t exhibit a variable pattern assciated with the grwth f sequential fllicular waves (fllicles grwing frm 3 t >5 mm in diameter) in individual ewes (6), it was nt pssible t develp a generalized divisin f the intervulatry perid int distinct phases f increased/decreased estradil prductin in all animals. Perid 1, r the phase f luteal tissue frmatin (crrespnding t the stage f the crpus hemrrhagicum), was regarded as the time after vulatin when CL were nt identifiable with ultrasngraphy (Days 0 t 2). Perid 3, r the phase f mid-cycle luteal functin, was defined as the perid encmpassing all the days n which dally mean serum cncentratins f prgesterne did nt differ significantly frm the maximal mean cncentratin f the hrmne recrded, in bth breeds, n Day 11; this perid lasted frm Day 7 t Day 12 in bth the breeds f sheep under study. The time between the end f luteal tissue frmatin and the beginning f the perid f mid-cycle luteal functin was designated Perid 2, r the phase f luteal grwth (Days 3 t 6). Perid 4, r the perid f luteal regressin and the fllicular phase, was the time frm the beginning flutelysis t the day befre vulatin. Fr all 4 perids abve, the relatinships between daily values f vaginal impedance and serum cncentratins festradil and prgesterne, and the E2:P 4 rati were determined by simple linear regressin. The independent variable was vaginal impedance while the dependent variable was serum cncentratin f estradil r prgesterne, r the E2:P 4 rati. Estrus Cycle RESULTS At the first bserved vulatin f the intervulatry interval studied, all the ewes but ne were well marked by rams, ne animal was prly marked, and at the secnd vulatin bserved, all ewes were well marked. The length f the intervulatry interval ranged frm 16 t 1 d in Western white-faced ewes (n = 10), and frm 16 t 17 d in Finn ewes (n = 7). The mean intervulatry interval was 17.0 ± 0.3 and 16.6 ± 0.2 d, fr Western white-faced and Finn ewes, respectively (P>0.05). Vaginal Mucus Impedance Dally serum cncentratins festradil and prgesterne, the E 2:P4 ratis and crrespnding values f vaginal mucus impedance are shwn fr 2 Western white-faced in Figure 1, and 2 Finn ewes in Figure 2. There was a significant main effect f day (P<0.001) but nt f breed (P = 0.9) while the day-by-breed interactin apprached significance (P>0.05) fr mean daily impedance f the vaginal mucsa in the Western white-faced and Finn ewes studied (Figure 3a). In Western

7 Therigenlgy 19 B 60 (a) 3" g 4 :c) 55 ~- 5 3.~ 45 ~ ],,3 40 ~ 35 ~ 30 ~ 25-2 l ' I ' U ' I ' I ' I ' I ' I ' I ' I O" t ] ~" 10 (b) 0 l (d) gl ~ } 100 ~ ~ ~ ' I ' W ~ F r I ' I ' I ' I ' I ' I ' I I I ~ F ~ ' ~ ~ - - r T ~ Days frm first vulatin f the intervulatry interval Figure 3. Mean (4- SEM) daily vaginal mucus impedance (a), serum cncentratins f estradil 0a) and prgesterne (c), and estradil:prgesterne ratis (d) nrmalized t the day f first vulatin f the intervulatry interval in 10 Western white-faced (11, A, -bld, *) and 7 Finn sheep ([3, A, -, 0) examined ver 1 vulatry cycle in the mid-breeding seasn (Octber t December). See text fr statistical details.

8 20 Therigenlgy white-faced ewes, mean vaginal impedance increased frm Day - 1 t Day 2 (P<0.05), remained abve 40 hms frm Day 2 t 14, and then declined (P < 0.05) t a minimal value n Day 16 after vulatin. Vaginal mucus impedance was <40 hms in all white-faced ewes n Days - 1 and 16 f the cycle, in 9 ut f 10 animals n Day 15, and in 3 ewes n each day f vulatin (at the beginning r end f the intervulatry perid studied). During the perid frm Day 2 t 14, vaginal mucus impedance decreased belw 40 hms in nly 1 ewe, n Days 10 and 12 t 14 f the estrus cycle (Figure lb). In Finn sheep, mean vaginal impedance increased frm a minimum n Day - 1 t Day 0 (P<0.05), it was variable but stayed abve 40 hms frm Day 0 t Day 13, decreased between Day 13 and Day 15 (P<0.05), and subsequently increased (P<0.05) between Day 15 and Day 17 f the cycle (Figure 3a). Values were <40 hms in all Finn ewes studied n Days - 1 and 14 t 15, in 1 ewe n Day 0 (day f the first vulatin f the intervulatry interval studied), in 2 ewes n Day 13, and in 4 ewes n Day 16 after vulatin. Vaginal mucus impedance during mid-cycle fell t <40 hms n nly 1 d (Day 11) in a single ewe (Figure 2b). Serum Cn~ntratins f Estradil and Prgesterne We previusly reprted (6) that transient increases in daily serum cncentratins festradil, with a peridicity f-4 d, were identified in all f the animals studied; the nset f these elevatins was assciated with the emergence f fllicular waves (fllicles grwing frm 3 t ~ 5 mm in size), and peak estradil cncentratins ccurred in the prximity f days n which the largest fllicles f sequential fllicular waves reached their maximum diameters. Peaks f transient increases in serum estradil cncentratins did nt vary (P>0.05) with the stage f the estrus cycle within breeds (means: 4.4 q- 0.2 and 6.7 q- 0.5 pg/ml fr Western white-faced and Finn ewes, respectively). There were significant main effects f day and breed (P<0.001) fr daily serum cncentratins f estradil nrmalized t the day f first vulatin, but their interactin was nt significant (P = 0.25). Mean daily serum cncentratins f estradil were significantly higher in Finn than in Western white-faced ewes n Days 2 t 5 and 14 t 15 after vulatin (Figure 3b). In this grup f Western white-faced ewes, mean daily serum prgesterne cncentratins rse frm Day 0 t reach a peak value n Day 11 f the cycle. Serum prgesterne cncentratins fell by Day 13 (P<0.05) and reached a nadir n Day 15 f the cycle. In the Finn ewes, daily mean serum prgesterne cncentratin increased t a maximum value n Day 11, then declined (P<0.05) by Day 13 and again n Day 14 (P<0.05); there was n significant difference (P>0.05) frm Day 7 t 12 f the cycle. Mean daily serum prgesterne cncentratins were higher (P<0.05) in Western white-faced than in Finn ewes frm Day 4 t 14 f the cycle (Figure 3c). Estradil:Prgesterne Rati There was a significant main effect f day (P<0.001), breed (P = 0.02), and a day-by-breed interactin (P = 0.04) fr daily values f the E2:P 4 rati ver Days - 1 t 17 when the data were nrmalized t the day f first vulatin f the intervulatry perid studied (Day 0; Figure 3d). In Western white-faced ewes, the E2:P 4 rati decreased significantly frm Day - 1 t Day 7 after vulatin and subsequently increased frm Day 12 t Day 14 (P<0.05). In Finn ewes, there was a

9 - l Therigenlgy 21 decline in the E2:P 4 rati frm Day 1 t Day 5 (P<0.05); the ratis n Days 5 t I0 and 12 did nt differ (P > 0.05), but the rati increased frm Day 11 t Day 14 (P<0.05) and subsequently declined frm Day 15 t Day 17 f the cycle (P<0.05). The mean daily values fr the E2:P 4 rati were significantly higher in Finn than in white-faced ewes n Days 3 t 10 and 12 t 15 after vulatin. Vaginal Mucus Impedance and Serum Estradil and Prgesterne Cncentratins Twards the End f the Cycle There was n significant main effect f breed (P = 0.54), but there was a significant main effect f day (P<0.001) and a day-by-breed interactin (P = 0.009) fr mean daily vaginal mucus impedance during the perid preceding the secnd vulatin f the intervulatry interval (Days - 5 t 0; Figure 4a). In Western white-faced ewes, vaginal impedance decreased (P<0.05) frm Day - 5 t Day - 1 befre vulatin and increased (P<0.05) n the day f vulatin (frm Day - I t Day 0). In Finn ewes, mean vaginal impedance declined sharply (P<0.05) frm Day -4 t Day -2, remained lw n the day preceding vulatin and increased (P<0.05) n Day 0. Daily serum cncentratins f estradil fr the perid f 5 d befre and n the day f vulatin differed significantly by day and amng breeds (P<0.001), and the day-by-breed interactin was als significant (P = 0.007; Figure 4b). In Western white-faced ewes, estradil secretin increased (P<0.05) frm Day -4 t Day -2 befre vulatin but did nt change significantly ver the next 2 d. Befre vulatin in Finn ewes, circulating cncentratins festradil increased frm Day -4 t Day -2 (P<0.05) and then declined t Day 0 (P<0.05). Finn sheep exceeded whitefaced ewes in mean daily serum cncentratins f estradil frm Day -4 t Day -2 prir t the secnd vulatin f the intervulatry interval studied (P<0.05). The analysis f mean daily serum cncentratins f prgesterne relative t the day f the secnd vulatin f the estrus cycle shwed a significant main effect f day (P<0.001) and breed (P = 0.003) but nt a day-by-breed interactin (P = 0.20; Figure 4c). Prgesterne cncentratins fell (P<0.05) frm Day -5 t basal r nndetectable levels n Day -2 prir t vulatin in bth breeds f sheep under study. Frm Day -5 t Day -2, prgesterne prductin by regressing CL was maintained at a higher level in white-faced than in Finn sheep (P<0.05). The E2:P 4 rati fr the terminal prtin f the estrus cycle differed between breeds (P = 0.02) and by day (P<0.001), and there was a significant breed-by-day interactin (P = 0.003; Figure 4d). E2:P 4 ratis were significantly higher in Finn than in white-faced ewes frm Day -3 t Day befre vulatin (Day 0). In white-faced ewes, the mean E2:P 4 rati increased (P<0.05) frm Day - 4 t Day - 2, but it did nt differ (P>0.05) ver the next 3 d (Day - 2 t Day 0). In Finn sheep, the E2:P 4 rati increased significantly ver the perid frm Day -5 t Day -2, and then fell (P<0.05) frm Day -2 t Day 0. The Relatinship between the Vaginal Mucus Impedance and Serum Cncentratins f Estradil and Prgesterne The results f regressin analyses fr daily serum cncentratins f estradil and prgesterne, and daily recrdings f vaginal mucus impedance during different stages f the

10 22 Therigenlgy I~ 60 (a) ~4 (c) 55 ~ 5 3 e~ 45 ~ 40 2 g ~ 35,~ 3 e~ "~ ~ ~" 10 (b). t (d) 0 l= 4 "~ [1 g 0-6 -T-- I " ---F I -T- I -I I "1 ] I-'---q Days frm secnd vulalin f the inlervulalry inlerval Figure4. Mean (+ SEM) daily vaginal mucus impedance (a), serum cncentratins f estradil (b) and prgesterne (e), and estradil:prgesterne ratis (d) fr the day f and fr 5 days prir t the secnd vulatin f the intervulatry interval, as determined in 1 0 Western white-faced 011, A, -bld, #) and 7 Finn sheep (D, zx, -, O) studied during the middle prtin f the breeding seasn (Octber t December). See text fr statistical descriptins.

11 Variables Perid t Perid 2 Perid 3 Perid 4 Days 0 t 2 a Days 3 t 6 Days 7 t 12 Days 13 t - 1 E 2 versus VMI r = 0.30, P = 0.11 r = -0.04, P = 0.2 r = -0.02, P = 0. r = -0.06, P = 0.70 r = -0.10, P = 0.67 r = 0.13, P = 0.50 r = 0.23, P = 0.1 r = -0.17, P = 0.42 r = 0.62, P = P4 = ~x VMI ~ r = 0.61, P = P4 = -t ~x VMI v r = -0.56, P = E2:P 4 = x VMI ~ r = -0.45, P = 0.03 E2:P 4 = ~x VMI v Oa IX).O Table 1. Crrelatins (with regressin equatins fr statistically significant crrelatins) between dally serum cncentratins f estradil (E 2, pg/ml), prgesterne (P4, ng/ml) r the estradil:prgesterne rati (E2:P 4, dependent variables), and vaginal mucus impedance (VMI, hms, independent variable). The data fr this analysis were btained fr 1 vulatry cycle in 10 Western white-faced (upper lines) and 7 Finn (lwer lines) ewes, and were divided int 4 different perids f the eslrus cycle based n the appearance f crpra lutea and prgesterne secretin. P4 versus VMI r = 0.61, P = P4 = { x VMI v r = 0.27, P = 0.23 r = -0.14, P = 0.39 r = 0.0, P = 0.53 r = 0.21, P = 0.27 r = 0.23, P = 0.14 E2:P 4 versus VMI r = -0.36, P = E2:P 4 = ~1 x VMI v r = -0.33, P = 0.14 r = , P>0.95 r = , P>0.95 r = -0.03, P = 0.6 r = 0.15, P = 0.35 a Day 0 = day f vulatin. b Lwer and upper 95% cnfidence interval limits.

12 24 Therigenlgy estrus cycle in ewes are summarized fr bth breeds in Table I. In Western white-faced ewes, there was n crrelatin between daily serum cncentratins f estradil and vaginal mucus impedance fr all 4 perids f the estrus cycle studied. Hwever, daily serum cncentratins f prgesterne were psitively and significantly crrelated with impedance f the vaginal mucsa during Perids 1 and 4 f analysis but nt during the phase f luteal grwth (Perid 2) r during the mid-luteal phase (Perid 3). Daily values fr the F_.2:P 4 rati were negatively and significantly crrelated with vaginal impedance during Perid 4 f the estrus cycle in white-faced sheep and during Perid 1 this crrelatin apprached significance. The difference between slpes f regressin equatins fr daily serum cncentratins f prgesterne (dependent variable) and vaginal impedance (independent variable) fr Perids 1 and 4 in Western white-faced ewes apprached significance (P = 0.07). The slpes f regressin lines fr the E2:P 4 rati and vaginal mucus impedance fr Perids 1 and 4 did nt differ (P = 0.13), and the linear relatinship between the 2 variables, fr bth perids cmbined, is given by the equatin: [Estradil (pg/ml):prgesterne (ng/ml)] = x [Vaginal Mucus Impedance (hms)]. Daily serum cncentratins festradit and prgesterne as well as the values fr the E2:P 4 rati were nt crrelated with vaginal mucus impedance during Perids 1, 2 and 3 f the estrus cycle in Finn sheep. There was, hwever, a significant crrelatin between daily serum cncentratins f prgesterne and vaginal impedance, and between daily E2:P 4 ratis and vaginal impedance during the terminal phase f the cycle (Perid 4). DISCUSSION During the fllicular phase f the estrus cycle in ewes, cervical mucus increases in amunt and becmes mre elastic (spinnbarkeit; 3). It has been suggested that these changes are due t the unppsed actin f estradil, as they d nt ccur during the luteal phase f the cycle (3, 5). Prgesterne priming in variectmized ewes results in higher levels f cervical secretin after injectin festradil (5). The ph fcervie-vaginal mucus in cyclic ewes declines rapidly ver the first 5 d f the cycle (Day 0 = day f estrus) and then mre slwly, t reach minimal values during the mid-luteal phase (between Days 10 and 13), after which it gradually increases (20, 26, 30). The chlride cntent f the vaginal mucus als varies thrughut the estrus cycle in sheep; NaC1 cncentratins ranged frm 0.1 t 0.2% n Days 1 t fllwing estrus, frm 0.2 t 0.5% n Days 9 t 14, and frm 0.7 t 0.9% during behaviral estrus (12). Ovulatin cincides with a marked increase in the mucsal NaC! cntent (12). The rate at which prductin fcervic-vaginal mucus, its physical prperties and inic cntent change acrss the estrus cycle may ultimately lead t alteratins in vaginal impedance. In the present study, there was a clse tempral relatinship between the decrease in vaginal mucus impedance belw 40 hms and manifestatins f behaviral estrus in all the ewes studied. These results are similar t previus bservatins in ewes (15, 2, 1, 9, 23, 1, 2, 29) and heifers (15, 24, 4,, 7, 27). It has been suggested that a marked decline in vaginal impedance reflects a rise in peripheral cncentratins festradil during the perivulatry perid. In variectmized animals, vaginal impedance remains elevated and des nt fluctuate (16), and treatment with exgenus estradil induces estrus behavir (14, 10). In the present study, there was n significant crrelatin between serum cncentratins festradil and impedance f the vaginal mucsa at any stage f the

13 Therigenlgy 25 vine estrus cycle. Alternatively, there was a psitive crrelatin between serum cncentratins f prgesterne and vaginal impedance late in the cycle in prlific Finn sheep, and during CL frmatin and lutelysis in nnprlific Western white-faced ewes. Additinally, daily E2:P 4 ratis were negatively crrelated with changes in vaginal mucus impedance in ewes during the perids abve. These results suggest that the electrical prperties f the vaginal mucsa f cyclic ewes change primarily due t alteratins in circulating cncentratins f prgesterne and/r due t an increase in the E2:P 4 rati. Hwever, based n a lack f crrelatin between estradil secretin and vaginal impedance in the present ewes, estradil des nt appear t exert a direct effect n impedance f the vaginal mucsa independent f prgesterne. Mean serum cncentratins festradil and prgesterne and the E2:P 4 rati differed during the mid-luteal phase f the estrus cycle between prlific Western white-faced and nnprlific Finn sheep, but there was n difference in mean daily values fr vaginal impedance. At lutelysis, the decline in circulating cncentratins f prgesterne t a basal level and the rise in estradil ccurred earlier in Finn cmpared t Western white-faced ewes. Serum prgesterne cncentratins declined at the same rate in bth breeds, whereas serum estradil cncentratins increased mre rapidly and- t a greater peak in Finn than in white-faced ewes. During the terminal phase f the cycle, vaginal mucus impedance decreased earlier in Finn than in white-faced ewes, but the nadir values did nt differ between breeds. Serum cncentratins f prgesterne, but nt f estradil, and the E2:P 4 rati were crrelated with vaginal mucus impedance late in the cycle but nt during the mid-luteal phase in the 2 breeds studied. Taken tgether, these results suggest that plgesterne levels typical f the mid-luteal phase maintain vaginal impedance in the face f varying serum cncentratins festradil and that a decline in serum cncentratins f prgesterne t lw levels, in the presence festradil, initiates a transient decrease in vaginal mucus impedance. In anther study, after the final luteal phase f the breeding seasn in ewes, there was a decline in vaginal mucus impedance, but there was n evident increase in circulating cncentratins festradil, and n ewe was marked by crayn-harnessed rams (unpublished data). The latter bservatin indicates that the decrease in serum prgesterne cncentratins induces a decline in vaginal impedance even in the absence f a nrmal fllicular phase level f estradil. In the present study, there was n linear relatinship between serum cncentratins f estradil r prgesterne and vaginal impedance during mid-cycle in ewes. We cncluded that impedmetrie prperties f the vaginal mucsa are f n value in estimating circulating cncentratins f the sterid hrmnes during the perid in questin. Vaginal mucus impedance des nt reflect the ttal vlume f luteal tissue and the presence r stage f develpment f the largest antral fllicles during the luteal phase f the vine estrus cycle prir t the nset f lutelysis. Hwever, during the terminal prtin f the intervulatry interval in sheep, frm apprximately Day 13 f the cycle until the next vulatin, vaginal impedance may serve as an accurate indicatr f serum cncentratins f prgesterne and E2:P 4 ratis. Thus, vaginal mucus impedance prvides useful infrmatin abut the curse f lutelysis and develpment f prevulatry antral fllicles in sheep. In summary, bielectrical impedance measurement f the vaginal mucsa, near the cervix uteri in ewes f different gentypes, is a useful tl fr the detectin f estrus and vulatin, and it als permits the estimatin f serum cncentratins f prgesterne and E2:P 4 ratis during

14 26 Therigenlgy lutelysis and the fllicular phase f the sheep estrus cycle. Vaginal mucus impedance appears t be primarily cntrlled by circulating cncentratins f prgesterne, but it als changes in respnse t shifts in the E2:P 4 rati when serum cncentratins f prgesterne are declining r are lw. REFERENCES 1. Abdel-Rahim SEA, el-nazier AE. Electric resistance f the vaginal mucus as an aid fr heat detectin in sheep. Vet Rec 197; 121: Adam L, Aizinbud E, Tadmr A, Schindler H. Impedmetrie prperties f the vulvar and vaginal tissues f ewes during the estrus cycle. J Reprd Fertil 191; 61: Adams NR, Tang BY. Changes in vine cervical mucus in respnse t estrgen treatment. J Reprd Fertil 1979; 57: Aizinbud E, Schindler H, Adam L. Impedmetry f the bvine vagina and vulva fr estrus detectin: present state f research and prspe~cts. Refuah Veterinarie 190; 37: Allisn AJ. Prductin f cervical mucus in ewes treated with exgenus prgestagen and estrgen. Austr J Agric Res 1972; 23: Bartlewski PM, Beard AP, Ck S J, Chandlia RK, Hnaramz A, Rawlings NC. Ovarian fllicular dynamics and their relatinships with endcrine variables thrughut the estrns cycle in breeds f sheep differing in prlificacy (J Reprd Fertil, in press). 7. Canfield RW, Butler WR. Accuracy f predicting the LH surge and ptimal inseminatin time in Hlstein heifers using a vaginal resistance prbe. Therigenlgy 199; 31: Carter PD, Duty JH. Assessment f vaginal impedance measurements as an indicatr f estrus in cattle. Austr Vet J 191; 56: Dumas G, Lemay JP. Measuring the cnductivity f vaginal mucus during estrus cycle in ewes. Medecin Veterinaire du Quebec 1990; 20: (in French with English abstract). 10. Yabre-Nys C, Martin GB, Venier G. Analysis f the hrmnal cntrl f the female sexual behavir and the prevulatry LH surge in the ewe: Rles f quantity f estradil and duratin f its presence. Hrmnes and Behavir 1993; 27: Hafez ESE. Functinal Anatmy f Reprductin in: Reprductin in Farm Animals. Philadelphia: Lea & Febiger, 197, p Hnmde J, Paehlag SV. Time f vulatin and chlride cntent f vaginal mucus during estrus in sheep. Indian J Anita Sci 1973; 43: Jseph IBJK, Currie WD, Rawlings NC. Effects f time after varieetmy, seasn and estradii n luteinizing hrmne and fllicle-stimulating hrmne secretin in variectmized ewes. J Reprd Fertil 1992; 94: Land RB, Thmpsn R, Baird DT. The duratin f estrus in varieetmized Finnish Landrace and Scttish Blackface ewes fllwing prgeterne and estrgen treatment. J Reprd Fertil 1972; 30: Leidl W, Stlla R. Measurement f electric resistance f the vaginal mucus as an aid fr heat detectin. Therigenlgy 1976; 6: Metzger E, Freytag R, Leidl W. Ger~it zur Messung der elektrisehen Leitfahigkeit des Vaginalsehleimes flit die Brunstfeststellung beim Rind. Zuchthyg 1972; 7:56-61 (in German).

15 Therigenlgy Murdch BE, O'Shea T. Activity f enzymes in the mucsal tissues and rinsings f the reprductive tract f the naturally cyclic ewe. Austr J Bil Sci 197; 31: Olic I, Rezac P, Pschl M. Impedance changes in the vaginal mucsa at the different stages festrus cycle in Muttn Merin ewes. Zivc Vyr 1990; 35: Ravindra JP. Gnadtrphins and Ovarian Functin in Ewes. PhD thesis, University f Saskatchewan, Rawal CVS. ph f vaginal mucus f Muzaffarnagari ewes during estrus. Indian J Anim Sci 195; 55: Rawlings NC, Jeffcate IA, Reiger DL. The influence festradil and prgesterne n peripheral serum cncentratins fluteinizing hrmne and fllicle stimulating hrmne in the varieetmized ewes. Therigenlgy 194; 22: Restall BJ. Histlgical bservatins n the reprductive tract f the ewe. Ausla" J Bil Sei 1966; 19: Rezae P, Olie I. Electrical prperties f vaginal and vestibular mucsa during the estrus cycle f ewes. Scientia Agrieultura Bhemslv 1990; 22: Schams D, Sehallenberger E, Hffmann B, Karg H. The estrus cycle f the cw: Hrmnal parameters and time relatinships cncerning estrus, vulatin, and electrical resistance f the vaginal mucus. Acta Enderinl 1977; 6: Sehrick FN, Surface RA, Prichard JY, Dailey RA, Twnsend EC, Inskeep EK. Ovarian structures during the estrus cycle and early pregnancy in ewes. Bil Reprd 1993; 49: Singh R, Rawal CVS, Kumar D. Use festrus cycle curve fph values f vaginal mucus t judge a sheep in estrus. Indian J Anim Sei 199; 59: Smith JW, Spahr SL, Pucker HB. Electrical cnductivity f reprductive tissue fr detectin festrus in dairy cws. J Dairy Sei 199; 72: Szezepanski W, Milewski S, Brzstwski H, Tanski Z, Czemiawska-Zajae S. Detectin f estrus and pregnancy in sheep basing n electrical resistance f vaginal mucus. Acta Aeademiae Agriculturae ae Teehnieae Olstenesis 1994; 41: (in Plish with English abstract). 29. Tekeli T, Aksy M, Cyan K, Alan M, Semaean A, Ayar A. Estrus detectin and estrus cycle patterns in sheep mnitred by electrical resistance. Livest Adviser 1992; 17: Zibi LO. Detectin f estrus in ewes by means f the acidity and elasticity feervievaginal mucus and vaginal temperature. Trpicultura 1990; :27-30 (in French with English abstract).