Prolactin in the Cat: II. Diurnal Patterns and Photoperiod Effects

BIOLOGY OF REPRODUCTION 28, 933-939 (1983) Prolactin in the Cat: II. Diurnal Patterns and Photoperiod Effects DONELLE R. BANKS and GEORGE H. STABENFELDT Department of Reproduction School of Veterinary Medicine University of California Davis, California 95616 ABSTRACT Photoperiod effects on prolactin (PrI) secretory patterns were investigated in the domestic cat. Circadian patterns of Pr! release were studied in the cat in blood samples obtained every 4 h. Plasma concentrations were significantly elevated during the dark period (X=31.7 ng/ml) compared to the light period (X=5.5 ng/ml) when blood samples were obtained under red light during the dark period. If however, a short period of exposure to white light occurred during the dark sampling time, significant differences were not observed. In a long-term study of the effect of photoperiod, animals subjected to a decreasing photoperiod (4 discrete changes from 14 h to 7 h light per day occurring over an 8-week period) showed no sgnificant difference in morning Prl concentrations during the 7-week period of 7 light per day (X-12.6 ± 0.3 ng/m!) compared to a group of cats maintained under 14 light per day throughout (X.17.2 ± 4.9 ng/ml). Three of 4 animals in decreased light did not have another estrous period following the initial light change (14 h to 12 h light); 1 animal had one esarous period following the photoperiod change. When the light was increased to 10 h per day, a return to estrus occurred within a period of 2 to 6 weeks. During the period of increased light, there was no change in PrI concentrations as compared to the decreased light period. These data indicate that major shifts in Pr! secretion do not accompany the seasonal anestrous period of the cat. The possibility does remain (based on the circadian data) that PrI secretion may be modified on a daily basis in response to photoperiod. INTRODUCTION The domestic cat is seasonally polyestrus, with cyclic ovarian activity occuring between January and September and ovarian inactivity from October through December (Scott and Lloyd-Jacob, 1959). Sheep exhibit a seasonal anestrus during which prolactin (Prl) concentrations are elevated during the period of ovarian inactivity (Munro et a!., 1980). Pr! levels have also been shown to be elevated during lactational anestrus in a number of species (sheep, Lamming et a!., 1974; goats, Hart, 1974; rats, Lu et a!., 1976). Furthermore, it has been demonstrated that hyperprolactinemia in women often results in amenorrhea (Franks et a!., 1975). Thus elevated Pr! has been found to be a factor common to several syndromes involving ovarian inactivity. On a more acute basis, many mammalian species Accepted January 6, 1983. Received September, 1982. Reprint requests: Donelle R. Banks, School of Veterinary Medicine, Dept. of Reproduction, Univemity of California, Davis, CA 95616. exhibit diurnal rhythms of Prl secretion that are controlled by light/dark cues. Rats and monkeys, for example, show elevated PrI concentrations during the dark period (Dunn et a!., 1972; Quadri and Spies, 1976). The cat is a useful model for comparative studies on photoperiod effects on ovarian cyclicity in that, as contrasted to sheep and goats, decreased photoperiod results in the cessation of ovarian activity. The present study was initiated to: 1) study the relationship of diurnal light changes on Pr! secretion in the cat and 2) study the effects of long-term photoperiod manipulation on Pr! secretion. Experiment MATERIALS AND METHODS I This experiment was designed to examine the diurnal PrI secretion patterns in the cat. The 8 domestic (mixed breed) cats used in this experiment had been maintained in 7 X 9 rooms under 14 h light and 10 h dark for several months. Lights were on from 0600 until 00 h. Cats were checked daily for estrus with a vasectomied male. None of the animals were sexually receptive on the day of the experiment. Blood samples were taken every 4 h for a 24-h period starting with 0800 h; the 933 Downloaded from https://academic.oup.com/biolreprod/article-abstract/28/4/933/27663

934 BANKS AND STABENFELDT sampling procedure has been previously described (Banks et al., 1983). The cats had previously been conditioned to the bleeding procedure, usually over a period of months, and normally did not exhibit any signs of emotional or physical stress. During the dark period, 4/8 cats were bled under a red light with no exposure to white light, while the other 4 cats were exposed to approximately 3 mm of white light during the sampling period. All samples were stored at -#{176}C until assayed. All samples were run in a single assay. PrI was determined (in both experiments) by radioimmunoassay (RIA) as previously described (Banks et al., 1983). Data were analyed by Student s t test or by an analysis of variance followed by ScheffC s method for multiple comparisons (Scheff, 1959). Experiment 2 This experiment was designed to test the effect of decreasing photoperiod on PrI concentrations during the fall and winter months and to test the effect of increasing photoperiod on Pd concentrations during the resumption of ovarian activity. Control animals were maintained under a photoperiod regimen compatible with continued cyclic ovarian activity (14L: 1OD) during the same period of time. Two groups of cats were housed in 7 X 9 rooms. One group of 6 cats was exposed to a lighting schedule of 14 h light in order to maintain normal cyclic ovarian activity. A second group of 6 cats was subjected to a regimen of decreased lighting as follows: 14L (10 weeks), 12L (2 weeks), 1OL (4 weeks), 8L (2 weeks), and 7L (14 weeks). This artificial lighting schedule resulted in the decreased photoperiod being extended for 7 weeks beyond the natural winter solstice (Dec. 21). The 7-h light period was increased on February 6 to 10 h light to stimulate ovarian activity. Both groups of cats were bled between 0800 and 0900 at least 3 times each week throughout the experimental period. Two cats from each group were dropped from the breeding schedule due to accidental matings resulting in pregnancy or pseudopregnancy. One cat was added to the decreased light group at the 3rd week of the 74i light regimen. This cat had been with the group from the beginning of the experiment but had been nursing kittens following an earlier pregnancy. All plasma samples were stored at -#{176}C until the PrI assay was performed. All samples from any one cat were run in a single assay. The time to onset of ovarian cyclicity from the initial increase in photoperiod was recorded. Experiment 1 RESULTS Figure 1 shows the changes in PrI concentrations over a 24-h period for the 2 groups of cats. For the group which had no exposure to white light during the dark bleeding periods (00, 2400 and 0400 h), Pr! concentrations increased approximately 10-fold between 1600 h (3.0 ± 0.2 ng/m!) and 00 h (29.7 ± 3.5 ng/ml). Lights had been off for 5 to mm 40 uj 0) +1 1( 2. I-. C.,.( 10#{149} -j 0 0. 0. #{212} 16 24 TIME OF DAY (14L: 100) FIG. 1. Plasma Pr! secretion levels in cats at 4-h intervals over a 24-h period. Lights were on from 0600 from 00 h; the dark period is designated by the dark area. #{149}-. represents cats (n=4) bled in the dark under a red light, o-o represents cats (n=4) exposed to approximately 3 mm of white light during the bleeding period. The initial 0800 h values for each group are indicated again in parentheses at the next 0800 h time. The vertical lines represent SEM s. prior to the 00-h blood samples. In this same group, Prl decreased approximately 3-fold between 0400 h (33.8 ± 3.5 ng/ml) and 0800 h (10.0 ± 0.7 ng/m!). Lights were on at 0600 h, or 2 h before the 0800 h bleeding. The group of cats which was maintained under the same lighting schedule (14L:1OD), but exposed to a few minutes of white light during the dark period bleedings, showed significantly different results (P<0.01). There was no significant rise in Pr! concentration during the dark hours in this group. Experiment 2 Figure 2 shows a composite of the Prl patterns in cats under decreased lighting and those maintained under a constant 14-h lighting schedule. The period of decreased lighting had no significant effect on daily Pr! concentrations. The average Prl levels during the 7-week period of 7 h light per day were similar to the subsequent 6-week period of 10 h light per day (13.2 ± 0.5 ng/m! vs. 12.6 ± 0.3 ng/ml, respectively). The,.#{231}ontrol group housed under 14 h light per day had values similar to the decreased photoperiod group during the equivalent 7-week period (17.2 ± 4.9 ng/ml vs. 13.2 ± 0.5 ng/m!, Downloaded from https://academic.oup.com/biolreprod/article-abstract/28/4/933/27663

DIURNAL AND ANESTROUS Prl IN CATS 935 l#{149}i! g 10 OM$TANT PH#{216}TOERIOO #{149} *4ANGINQ PWOTotMC0-0-0-. SEPTEMBER OCTOBER 15 1 15 1 15 NOVEMBER DEMSER JANUARY FEBRUARY 15 MARCH TIME OF YEAR FIG. 2. Plasma Pr! levels for two groups of cats subjected to 2 different photoperiod regimens. Group 1 (n-4) (.-.) was maintained under 14 h light per day throughout the experimental period. Group 2 (n=4) (a-o) was subjected to a changing photoperiod as indicated by the hours of light shown along the abscissa. For clarity, the SEM s are not shown; the 2 groups are not significantly different (P0.18). P0.18). Two individuals in the 14 h group had consistently higher Pri concentrations throughout the entire period. Figure 3 shows individual PrI patterns for 4 cats throughout the 29-week experimental period of d#{231}creased photoperiod. No significan change in Pr! concentration was observed following the decrease in exposure to light. Following the increase from 7 to 10 h light per day there was a slight, but nonsignificant decrease in Pr! concentration. Figure 4 shows individual profiles of cats maintained in 14 h light per day. Two of the cats had large daily fluctuations which were significantly greater (P<0.05) than the variance observed for cats in the decreasing photoperiod regimen. The reason for the larger variance of this group is not known. Three of 4 animals in decreased light did not have another estrous period following the initial light change of 14 h to 12 h light; 1 animal had one estrous period following the photoperiod change. Following the increase of light from 7 to 10 h per day on February 6, there was a return to estrus in 4/5 cats within 60 days. The 4 cats commenced estrus at 14, 14, 29 and 38 days after the increase of light. Animals in the 14-h light group had continuous cyclic ovarian activity throughout the entire period as judged by periodic manifestations of estrus. DISCUSSION There was a consistent elevation of PrI secretion during the dark period (00, 2400 and 0400 h) in the 4 cats which were bled under a red light. At the time of the 00-h blood sample, the cats had been in darkness for only about 10 to 15 mm. Thus there appears to be a rapid release of Pr! within minutes following the onset of darkness. In the second group of cats who were exposed to a few minutes of light during each bleeding, the nocturnal elevation of Pr! was not observed, It appears that the brief exposure to light negates the diurnal Pr! rhythm that was established in the first group, although it is possible that Pr! secretory patterns could have changed between sampling periods. Several other species exhibit diurnal Prl rhythms. In rats, Dunn et al. (1972) noted that peak Pri levels were attained at h (5 h after lights were turned off) and lowest levels occurred at 0800 h (4 h after lights were turned on). These Prl changes suggest a lightcued response for the rat, similar to the relationship observed between Pr! and light in the cat. In our study, PrI was significantly lower at 0800 than 0400 h but decreased even further at 10 h (6 h after lights on). Pr! levels obtained between 0800 and 0900 h in the long-term photoperiod study are comparable to those found at the same time of day in the circadian study, a time of day when Pr! secretory patterns are in downward transition. It is likely that some of the variation observed in the long-term photoperiod study was due to 1) the fact that cats were sampled at a time of day when PrI concentrations were declining and 2) because there was often about an hour s variation in sampling time from day to day. In humans, there are marked increases in Pr! concentrations during sleep periods with Downloaded from https://academic.oup.com/biolreprod/article-abstract/28/4/933/27663

936 BANKS AND STABENFELDT maximal levels reached during the early morning hours. When sleep onset was delayed for 9 h, Prl elevation still occurred during sleep indicating a positive correlation with sleep and not darkness (Sassin et a!., 1973). Monkeys sampled every 4 h showed an elevation at 2100 h (1 h after dark) which continued through the night with Prl concentrations being significantly lower during the day (Quadri and Spies, 1976). It is not known, however, if sleep patterns played a role in the nightime Pr! elevation in the simian study. In view of the sporadic sleep patterns of cats, it is difficult to determine if a relationship exists between Pr! secretion and sleep. The cats in this study were not observed continuously and thus it is not possible to relate the individual Pr! concentration changes with sleep-awake patterns. The Pr! release patterns in the cats in the chronic photoperiod portion of this study did not show a correlation with changes in photoperiod, either immediate or delayed. The transient decreases to slightly below average Pr! concentrations observed in 3 animals following CHANGING PHOTOPERIOD *63 I 10 40 #{149}*. *39 10#{149} *37 10 1 15 1 15 1 15 1 15 SEPT OCT NOV DEC 1 15 1 15 JAN FEB 15 MAR TIME OF YEAR FIG. 3. PrI levels in cats exposed to a short photopermod. The arrows indicate the time of the light change; the numbers represent the hours of light exposure. Downloaded from https://academic.oup.com/biolreprod/article-abstract/28/4/933/27663

D1URNAL AND ANESTROUS Prl IN CATS 937 CONSTANT PHOTOPERIOD 40 69 10. I a 70 60 *68 50-40 E 1 10 10 40 10 1 15 1 15 1 15 1 SEPT OCT NOV DEC TIME OF YEAR 15 JAN FEB MAR FIG. 4. PrI levels in cats maintained in a constant photoperiod of 14L:1OD. Downloaded from https://academic.oup.com/biolreprod/article-abstract/28/4/933/27663

938 BANKS AND STABENFELDT photoperiod change occurred at different time periods which probably reflect random fluctuations or transient responses to decreased photoperiod. This contrasts with data obtained in sheep wherein Prl concentrations were high during anestrus and low during the breeding season (Munro et a!., 1980). Since Prl is elevated during the dark hours in the cat, there would be a progressive increase in the duration of high concentrations of Pr! as light was shortened from 14 h to 7 h. It is therefore possible that the longer exposure to the elevated nocturnal Prl concentrations on a daily basis could suppress ovarian activity, a finding which could not be determined in the present study. The pineal is thought to influence ovarian activity, in a negative sense, through the production of melatonin. Our laboratory has recently developed evidence for cats (H. Leyva, unpublished observations) that a reduced period of darkness results in a lower production of melatonin. The decreased melatonin production in cats presumably allows gonadotropin secretion to be reestablished. Sheep are inhibited, in contrast to the cat, by increasing light. The importance of the timing of onset of light during the 24-h period explains this phenomenon as shown by Lincoln and Short (1980). They found that melatonin was released in greater amounts in the ram if darkness occurred at 2400 h as oposed to 1600 h onset (lights on again at 0800 in both cases). In this instance of a short-day breeder, a reduced period of darkness resulted in greater melatonin production and, conversely, a longer period of darkness coupled with an earlier onset of dark within the day resulted in lower melatonin production. In our study, the increase in light to 10 h in the decreased photoperiod group, while not affecting basal PrI during the lighting period did allow gonadotropin synthesis and release to increase which resulted in a return to estrus in 4/5 cats within 6 weeks. The variability of the elapsed time between the increased photoperiod and the first estrus (range, 14 to 38 days) demonstrates a large degree of variation among individual cats in responding to an acute stimulus. The response time for the reestablishment of ovarian activity under natural photoperiod following the winter solstice is not different from the response time obtained in this study by an acute light change. The cat resumes ovarian activity sooner following the lengthening photoperiod compared to sheep and horses. The average interval between winter solstice and the reestablishment of ovarian cyclicity is approximately days for the cat compared to 40-50 days (from the summer solstice) for sheep and 60-90 days (from the winter solstice) in horses. The cat provides a useful model for further investigation into the mechanisms by which photoperiod changes result in alternating suppression and stimulation of ovarian acitvity. The results may provide clues for the development of feasible means of contraception in the cat by the suppression of ovarian activity. The results could also provide clues for understanding the induction of ovarian activity in other more economically important species (e.g. sheep and goats). ACKNOWLEDGMENTS We thank Sharon Zito, Mary Kramer, Janet Lawson and Dr. Hugo Leyva for their assistance in the collection of blood samples and animal care, and Dr. Dennis Stewart for statistical assistance. We thank Dr. A. F. Parlow and NIAMDD for supplying canine prolactin RIA kit. This work was supported, in part, by the Animal Protection Institute, Sacramento, CA. REFERENCES Banks, D. R., Paape, S. R. and Stabenfeldt, G. H. (1983). Prolactin in the cat: I. Pseudopregnancy, pregnancy and lactation. BioL Reprod. 28: 923-93 2. Dunn, J. D., Arimura, A. and Scheving, L. E. (1972). Effect of stress on circadian periodicity in serum LII and prolactin concentration. Endocrinology 90:29-33. Franks, S., Murray, M.A.F., Jequier, A. M., Steele, S. J., Nabarro, J.D.N. and Jacobs, H. 5. (1975). Incidence and significance of hyperprolactinemia in women with amenorrhoea. CIin. Endocrinol. 4:597-607. Hart, 1. C. (1974). The relationship between lactation and the release of prolactin and growth hormone in the goat. In: Journal Reproductive Fertility Symposium Report No. 4 (J. S. Perry, ed.). Blackwell Scientific PubI., Oxford, London, p. 485. Lamming, G. E., Moseley, S R. and McNeilly, J. R. (1974). Prolactin release in the sheep. J. Reprod. Fertil. 40:151-168. Lincoln, G. A. and Short, R. V. (1980). Seasonal breeding: Nature s contraceptive. Recent Prog. Horm. Res. 36:1-43. Lu, K. H., Chen, H. T., Huang, H. H, Grandison, L., Marshall, S. and Meites, J. (1976). Relation between prolactin and gonadotrophin secretion in post-partum lactating rats. J. Endocrinol. 68:241-250. Munro, C. J., McNatty, K. P. and Renshaw, L. (1980). Circa-annual rhythms of prolactin secretion in Downloaded from https://academic.oup.com/biolreprod/article-abstract/28/4/933/27663

DIURNAL AND ANESTROUS Pr! IN CATS 939 ewes and the effect of pinealectomy. J. Endocrinol. 84:83-89. Quadri, S. K. and Spies, H. G. (1976). Cyclic and diurnal patterns of serum prolactin in the Rhesus monkey. Biol. Reprod. 14:495-501. Sassin, J. F., Frant, A. G., Kapen, S. and Weitman, E. D. (1973). The nocturnal rise of human prolactin is dependent on sleep. J. Clin. Endocrinol. Metab. 37:436-440. Scheff#{233},Henry. (1959). The Analysis of Variance. John Wiley & Sons, New York. Scott, P. P. and Lloyd-Jacob, M. A. (1959). Reduction in the anoestrous period of laboratory cats by increased illumination. Nature 184:22. Downloaded from https://academic.oup.com/biolreprod/article-abstract/28/4/933/27663