Relationship between body condition and levels of serum luteinizing hormone in postpartum mares

THERIOGENOLOGY

RELATIONSHIP BETWEEN BODY CONDiTION AND LEVELS OF SERUM LUTEINTZING HORMONE IN POSTPARTUM MARES 3 K.K. Hines,' S.L. Hedge,*3L . . Kreider, G.D. Potter and P.G. Harms Texas Agricultural Experiment Station4 Texas A & M University College Station, TX 77843

Received for publication: September 18, 1986 Accepted: September 25, 1987

ABSTRACT To study the endocrine environment of foaling mares at different levels of body condition, 12 mares were divided equally into two groups, one designed to foal at a condition score of 4.5 or less (thin) and the other to maintain a condition score of 6 or greater (control). Serum samples were collected twice weekly before foaling, and daily thereafter until 7 d following the second estrous cycle after parturition. Serum levels of luteinizing hormone (LH) were measured by Radioimmunoassay. In addition, mares were weighed, body condition was scored and body fat content was measured weekly. One consequence of low body condition at foaling in thin mares was a longer mean gestation length than in control mares. Also, the pattern of LH observed during the first and second estrous cycles after foaling was different for thin vs control mares. Serum LH concentrations for thin mares were greater during their second estrous cycle than during their first estrous cycle. Also, in thin mares, the interval from parturition to ovulation, as indicated by the day of maximum concentration of LH during estrus, was random. Control mares had equivalent concentrations of serum LH in both cycles, and they had predictable intervals from parturition to ovulation. 1-Hpatterns in thin mares were similar to those reported in mares during eeasonal transition, whereas LH patterns of control mares were similar to those reported in mares during the middle of the breeding season. Key words:

equine, body condition, LH, reproductive efficiency

Technical Article 21665, Texas Agricultural Experiment Station. The authors gratefully acknowledge the assistance of Dr. C.E. Gates, Institute of Statistics, Texas A&M Univ. in the statistical analysis of the data. 1 Present address: Department of Veterinary Science, University of 2Kentucky, Lexington, KY. Present address: Department of Animal Science, University of Florida, 3Gair.esville,FL. 4Present address: Dean Lee Research Station, Aiexandria, LA. Equine Research Program, Department of Animal Science.

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INTRODUCTION The interaction between nutrition and the resultant body condition and reproductive performance 3s of critical importance to management of the broodmare. A relationship between low body condition and reproductive inefficiency has been observed in many species, including cattle (1,2), humans (3.4) and horses (5-7). These reports generally confirm that poor body condition resulting from inadequate nutrition adversely affects reproductive performance. The aspects of reproductive performance which are adversely affected include pregnancy rates (2,5), length of postpartum anestrus (2), seasonal transition into the breeding season (6), onset of puberty (24,25) and other endocrine mechanisms which control reproduction (1,4). The interrelationships among the hormones of reproduction, particularly follicle stimulating hormone (FSH) and LH, are less understood in the mare than in other species. This is especially true of FSH (8). More is known regarding the role of LH in equine reproduction, but the effects of inadequate nutrition and lower body fat levels on this hormone are not understood. In contrast to other species, the mare has an early, fertile postpartum estrus which generally occurs within 12 d of foaling (8). Normally, concentrations of LH during this estrus are comparable to those seen at mid-breeding season (9,lO). However, in nonfoaling mares, the first estrous cycle of the breeding season is characterized by lower LH levels than during subsequent cycles (8,ll). The objectives of the current study were to examine the levels of LH in mares of high or low body condition during the late period, through foaling and the first and second estrous cycles. Secondly, the effects of body condition on reproductive performance were evaluated and related to levels of LH.

MATERIALS AND METHODS Twelve registered American Quarter Horse mares were selected from the broodmare band (n = 56) at Texas A&M University and divided into two treatment groups. Mares were divided into groups according to mean age and expected foaling dates (10 ?r 1.5 yr vs 10.5 + 1.9 yr and March 31 + 7 d vs April 2 + 6 d, for thin and control mares, respectively, P >0.5). Expected foaling date was determined by using a gestation length of 340 d from the time of conception. Treatment began the last week in January 1983, 48 d before the first mare foaled, and continued through August 1983, when the study ended. Mares in the thin group were fed to reduce body condition to a score of 4 or less prior to foaling (6). The diet for the thin mares was manipulated such that they received approximately 85% of the NRC recommended levels of protein and energy during the last 90 d of gestation. Gill et al. (20) have demonstrated that protein restriction of 70% of the NRC recommendation during the late gestation and lactatjon had no

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detrimental effects on reproduction, including plasma LH concentrations. Gill et al. (20) suggest that the NRC requirements for protein for late gestation in mares are high. Thus, the level of protein restriction prior to parturition in our study was considered to have no detrimental effects. Mares in the control group were maintained at a condition of 6 or greater through gestation and lactation (6), receiving NRC recommended levels of all nutrients. After parturition all mares received the NRC recommended levels of protein and energy, based on their mature body ,weight. Each week, beginning the week of February 25, mares were weighed, their body condition was scored and their body fat content was determined. Condition scores were assigned to the mares by three independent appraisers (6). Scores ranged from 1 to 9, with 1 representing extremely emaciated and 9 representing extremely fat animals. Body fat content was estimated by the method of Westervelt et al. (13) which correlates rump fat thickness with total extractable body fat. Rump fat thickness 5 cm from the midline and 10 cm from the point of the hip was measured by ultrasonic scanning (13). All mares were assigned condition scores, weighed and fat scanned on the same day. Beginning on Day 3 after foaling, all mares were teased with a vigorous stallion. Follicular development was monitored by palpation per rectum every second day at the onset of behavioral estrus, or 7 d post foaling if no behavioral signs had been observed. Mates were inseminated during their second postpartum estrus with 500x10 motile spermatozoa and were maintained in their treatment groups for 30 d follcswingconception to determine the incidence of early embryonic loss. Early indications of pregnancy included increased uterine tone, closed and pale cervix, and presence of fetal bulge (8). Serum LH was measured by RIA, validated for horse serum by Nett et al. (14) and modified by Hodge et al. (15). The first antibody, rabbit antiovine LH (GDN 615; 16) and the second antibody, sheep antirabbit gamma globulin (Antibodies Inc., Davis CA.) were titrated such that approximately 30 to 35% of the 125 I-labeled ovine LH (OLH-LER 1056-2) could bind in the absence of any competing LH. Highly purified equine LH (prep !/e98a,3xNIH-LH-Sl, gift from Dr. H. Papkoff: 17), was used as the :referencestandard. The range of the standard curve was from 5 ng eLH/tube to 20 pg eLH/tube. Intraassay variation was 5% (n = 8) and interassay variation ranged from 12% (low pool, n = 8) to 15% (high pool,,n = 9). Blood samples were collected by jugular puncture twice weekly before parturition and daily thereafter until 7 d following the second estrous cycle. The blood was allowed to clot for 20 h at 4" C, and serum was harvested following centrifugation. Serum was stored at -20°C until assayed. Serum pools were collected from an early esters mare (low poolj and from a second mare one day following ovulation (high pool).

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One measure of how much LH was released during estrus was the peak value of LH. Also the amount of LH released during estrus was estimated by calculating the area enclosed by the surge of LH from 4 d before the day of peak LH to 4 d following peak LH release. Both the peak level of LH and the area under the LH curve at estrus were corrected for levels of LH during diestrus, as these were different among individuals and between groups (0.47 + 0.3 ng/ml vs 0.75 f 0.1 ng/ml, thin vs control groups P
RESULTS The 12 mares selected for this study were placed in their treatment groups in January 1983. Initial physical measurements were made at the end of February. At this time, body weights were not different between the thin group and the control group (605 f 13 kg vs 626 + 20 kg, P>O.3). However, condition scores of the thin group were lower than scores of the control group (5.5 f 0.6 vs 6.8 + 0.7, P 0.4); however, thin mares had less body fat at foaling than control mares (P O.5). control mares was within the expected range (343 + 2.3 d, P>O.l), but the thin mares carried their foals significantly longer (352 +- 1.6 d, ? O.5). All of the thin mares conceived 45.6 2 4.7 d after foaling. Five of six control mares conceived and had an interval from foaling to conception of 48.6 _+ 8 d. The one mare that did not conceive was diagnosed as having an intrauterine infection. After antibiotic treatment, this mare conceived during the following breeding season. All mares delivered healthy foals the following spring. While estrous-cycle length was not affected by treatment (data not shown),the regularity of the first two cycles was influenced by the body condition of the mares (Table 1). In control mares, intervals from

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650 -

-G

-

Conml

-

Thin

Mares

Mares

Y

E

600

w ._ f C

cu

550 -

I” 500'

I

I

-4

1

t

-2

1

I

I

1

2

t

a

4

*

I

6

1

1

6

J

10

Foaling

Weeks From Foaling

Figurel.

&hanges in lxxiyweight, condition score and body fat percentage in control and thin mares. Dashed line represents the week of foaling. Bars represent one standard error.

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foaling to day of peak LH release during either the first (Cycle 1) or second postpartum estrus (Cycle 2) became shorter as daylength increased. When foaling date of control mares was used as an indicator of daylength (i.e. early date = short days, late date = long days), the decrease in days from foaling to LH peak during estrus best fits a simple linear model with negative slope and correlation. However, the same simple linear model does not fit the relationship between estrus and photoperiod in thin mares. Instead, the thin mares exhibited a random onset of estrus following parturition, as indicated by nonsignificant values for slope and correlation coefficient for Cycle 1 and cycle 2. Table 1.

Regression of Day of peak LH secretion during estrus, as a function of foaling date

Slope Cycle 2 Cycle 1

R Cycle 1

Cycle 2

Control mares

-0.16+

-0.20*

-0.80*

-0.83*

Thin mares

-0.10

-0.04

-0.40

-0.11

X(P< 0.1). (PC 0.5).

Patterns of LH in serum as measured by RIA showed the typical pattern (8). Generally, levels of LH were at their lowest levels before and after parturition and during diestrus. All mares exhibited the broad LH surge at estrus which is characteristic of the equine estrous cycle (Figure 2). Similar amounts of LH were released during estrus in Cycle 1 and in Cycle 2 (Table 2). Even though the difference between the maximum level of LH between Cycle 1 and Cycle 2 approached significance (PC O.l), the total amount of LH released during estrus in each cyclr ~3s similar (P>O.2). Thin mares, on the other hand, released more LH during the second rather than the first postpartum estrus. This is shown by both higher peak levels of LH during estrus of Cycle 2 as well as greater total LH release during estrus (PC 0.001).

DISCUSSION Mares were in their treatment groups from 48 to 93 d before parturition. Therefore, some thin mares were subject to restricted feed intake longer than others in the group. Even though later foaling mares in the thin group were fed a restricted diet longer, the treatment goal was to attain a body condition at foaling of less than 4.5. Previous work had demonstrated that protein restriction during both gestation and lactation had no effect on reproductive performance of mares (19,20).

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7

5_

CONTROL THIN

HARE

o--o

HARE

-

5-

L-

3-

,

r

Figure

2.

Levels of Lll in serun frcm one representative mare nay WI-CJ (day of foaling) fmboth frcm each grmlp . individuals is May 4.

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Table 2.

Serum LH during the LH surge at estrus

Cycle 1

Cycle 2

Control mares Peak LH Total LHa

4.0 f 0.4 18.1 f 1.7

6.1 f 1.0 23.0 2 4.0

Thin mares Peak LH Total LHa

3.5 f 1.4 15.7 f 4.1

7.4 f 3.0 30.7 + 5.9

Group

Difference

2.0 If0.8 4.9 ? 3.5 *** 3.9 + 0.6,,, 15.1 f 2.5

aTotal LH measured by estimation of area under the curve (peak day r 4 d) by Simpson's Rule. ,..*Valuesare means (n = 6) + SEM. (P< 0.001).

Furthermore, differences in weight and body fat content were not present between treatment groups as close as 4 wk before parturition. Also, the treatment effects discussed below were consistent within the thin-mare group. Therefore, length of time mares were on a restricted diet One of the apparently had minimal effects following parturition. effects of prefoaling treatment was an increased gestation length of thin mares. This observation is not consistent with recent research which showed no influence of either protein or energy restriction on gestation length in mares (5,19,21). However, Howell and Rollins (22) indicated that level of nutrition during gestation, in addition to photoperiod, can affect gestation length in mares. More recent work by Hodge et al. (15) supports the theory that photoperiod may be the dominant factor of gestation length. However, nutritional state may interact with photoperiod in influencing gestation length and it cannot be disregarded. Patterns of LH release in serum were used jn this study to indicate the incidence of estrous in these 12 mares. Thus, interpretation of behavioral intensity and dependence on manual palpation of the reproductive tract to specifically determine the stage and timing of the estrous cycle were avoided. The two groups of mares differed in the regularity of estrous cycles following parturition. Control mares demonstrated a predictable reduction in the interval between foaling and day of peak LH release for both Cycle 1 and Cycle 2 as daylength increased. Thin mares, in contrast, did not appear to respond to increased natural photoperiod, exhibiting no correlation between time of year and interval from foaling to either the first or second estrous cycle. This occurred even though thin mares were being fed adequate amounts of energy and protein after foaling to meet their maintenance and lactational needs.

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At the onset of the breeding season, LH released in mares is less than later in the breeding season (11). Early estrous cycles are characterized by LH-release patterns that are smaller in magnitude than LH-release patterns of estrous cycles in midbreeding season. The maximum amount of LH released during estrus appears to increase and reach a steady level as summer approaches. At the end of the breeding season, the highest observed value for LH concentration during the last estrous cycle of the year was lower than the cycle that immediately preceded it (23). T'hetwo groups of mares in this study exhibited different LH release patterns during Cycle 1 and Cycle 2. In control mares, the total amount of LH released during the estrous surge and the peak amount of LH relea,sedduring estrus were not different between the first and second cycle.3. This pattern of LH release was similar to that of normally cycling mares at midbreeding season (8). In contrast, thin mares relea,sedmore LP and had higher peak LH levels during the second cycle than during the first. This change in the levels of LH observed in consecutive estrous cycles in the mare is similar to the pattern observed in mares during transitions into and out of the breeding seaso>l. Even though reduced reproductive efficiency, as evidenced by pregnancy rate and interval from foaling to conception, was not observed in these six multiparous mares, previous work utilizing over 900 mares demonstrated the adverse effects of low body condition (< 4.5) at the onset of the breeding season (5,6). Altered reproductive function in thin mares in our study was characterized by an apparent asyncrony of estrous cycles with photoperiod and by altered endocrine patterns. Thus, these data indicate that effects of undernutrition, resulting in low body condition/low body fat content, may occur through impaired function of the brain-hypothalamic-pituitary axis and its control of This observation is consistent with recent reproductive hormones. findings that undernutrition may impair gonadotropin releasing hormone (GnRH) secretion, thereby interrupting normal reproductive function (24,2.5). It remains to be determined how undernutrition and/or low body fat signal the neural centers involved in controlling the release of the hormones involved in reproduction.

REFERENCES 1. Rutter, L.M. and Randel, R.D. Postpartum nutrient intake and body condition: Effect on pituitary function and onset of estrus in beef cattle. J. Anim. Sci. -58~265-274 (1984). 2. Dunn, T.G. and Kaltenback, C. Nutrition and the postpartum interval of the ewe, sow and cow. J. Anim. Sci. -51 (Suppl. II):29-39 (1980).

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3. Frisch, R.E. Pubertal adipose tissue: Is it necessary for normal sexual maturation? Evidence from the rat and human female. Fed. Proc. -39:2395-2400 (1980). 4. Warren, M.P. Effects of undernutrition on reproductive function in the human. Endocrine Reviews &:363-377 (1983). 5. Henneke, D.R., Potter, G.D. and Kreider, J.L. Body condition during pregnancy and lactation and reproductive efficiency of mares. Theriogenology -21:897-909 (1984). 6. Henneke, D.R., Potter, G.D., Kreider, J.L. and Yeats, B.F. Relationship between physical measurements and body fat percentage in mares. Equine. Vet. J. -15:371-372 (1983). 7. Day, F.T. breeding.

Some observations on the cause of infertility in horse Vet. Rec. -51:581-587 (1939).

8. Ginther, O.J. Reproductive biology of the mare: Basic and applied aspects. McNaughton and Gunn, Inc., Ann Arbor, MI, 1979, pp. 169-174. 9. Fitzgerald, B.P., I'Anson, H., Legan, S.J. and Loy, R.G. Changes in patterns of luteinizing hormone secretion before and after the first ovulation on the postpartum mare. Biol. of Reprod. -33:316323 (1985). 10. Loy, R.G., Evans, M.J., Pemstein, R. and Taylor, T.B. Effects of injected ovarian steroids on reproductive patterns and performance on postpartum mares. J. Reprod. Fertil. -32 (Suppl.):199-204 (1982) 11. Freedman, L.J., Garcia, M.C. and Ginther, O.J. Influence of photoperiod and ovaries on seasonal reproductive activity in mares. Biol. of Reprod. -20~567-574 (1979). 12. National Research Council. Nutrient requirements of horses 66. National Academy of Sciences, National Research Council, washington, D-C,, 1978. 13. Westervelt, R.G., Stouffer, J.R., Hintz, H.F. and Schryver, H.F. Estimating fatness in horses and ponies. J. Anim. Sci. -43~781-785 (1976). 14. Nett, T.M., Holtan, D.W. and Estergreen, V.L. Levels of LH, prolactin and estrogens in the serum of postpartum mares. J. Reprod. Fertil. -23 (Suppl.):201-206 (1975). 15. Hodge, S.L., Kreider, J.L., Potter, G.D., Harms, P.G. and Fleeger, J.L. Influence of photoperiod on the pregnant and postpartum mare. Am. J. Vet. Res. -43:1752-1755 (1982).

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Niswender, G.D., Midgley, A.R., Jr., Monroe, S.E. and Reichert, L.E. Radioimmunoassay for rat luteinizing hormone with antiovine LH serum and ovine LH-131 I. Proc. Sot. Exp. Biol. and Med. -128:807811 (1968). 17. Licht, P.A., Bona Gallo, A., Agarwal, B.B., Farmer, S.W., Castelino, J.B. and Papkoff, H. Biological and binding activities of equine pituitary gonadotropins. J. Endocrinol. -83:311-322 (1979). 18. !;teel,R.G.D. and Torrie, J.H. Principles and procedures of statistics - With special reference to the biological sciences. McGraw Hill Book Co., New York, 1960, pp. 161-251. 19. Gill, R.J., Potter, G.D., Kreider, J.L., Schelling, G.T. and .Jenkins,W.L. Postpartum reproductive performance of mares fed various levels of protein. Proc. 8th Equine Nutrition and I?hysiologySymposium. Lexington,KY. pp. 311-316 (1983j. 20. Gill, R.J., Potter, G.D., Kreider, J.L., Schelling, G.T., Jenkins, W.L. and Hines, K.K. Nitrogen status and postpartum LH levels of mares fed varying levels of protein. Proc. 9th Equine Nutrition and Physiology Symposium. East Lansing,MI, pp. 84-89 (1985). 21. 13anach,M.A. and Evans, J.W. Effects of inadequate energy during gestation and lactation on the estrous cycle and conception rate of mares and on their foal weights. Proc. 7th Equine Nutrition and :?hysiologySymposium. Warrenton,VA, pp. 97-100 (1981). 22. Howell, C.E. and Rollins, W.C. Environmental sources of variation tonthe gestation length of the horse. J. Anim. Sci. -10:789-805 (1951). 23. #Snyder,D.A., Turner, D.D., Miller, K.F., Garcia, M.C. and Ginther, 13.J. Follicular and gonadotrophic changes during transition from (ovulatoryto anovulatory seasons. J. Reprod. Fertil. -27 (supp1.):95-101 (1979). 24. Foster, D.L. and Olster, D.H. Effect of restricted nutrition on puberty in the lamb: Patterns of tonic luteinizing hormone (LH) secretion and competency of the LH surge system. Endocrinology .-116:375-381 (1985). 25. Bronson, recovery recovery hormone.

F.H. Food-restricted, prepubertal, female rats: Rapid of lutelnizing hormone pulsing with excess food and full of pubertal development with gonadotropin-releasing Endocrinology -118:2483-2487 (1986).

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