Search

The object of this study was to examine ovarian regulation of pulsatile luteinizing hormone (LH) secretion during early gestation. This was done primarily by analyzing pulsatile LH release in rats that were either sham ovariectomized (OVX) on Day 7 of pregnancy, implanted with empty Silastic capsules, and bled on Day 8, or OVX on Day 7, immediately implanted with Silastic capsules producing plasma levels of estradiol and/or progesterone characteristic of Day 7 to 8 of pregnancy, and bled on Day 8. In addition, the role of progesterone in regulating pulsatile LH secretion was also examined by administration of the progesterone receptor antagonist, RU486, on Day 7 and examining pulsatile LH release on Day 8 of pregnancy. OVX caused a marked increase in LH pulse amplitude and frequency within 24 h. Replacement with physiological plasma levels of estradiol or progesterone alone had no suppressive effect on this OVX-induced increase in pulsatile LH secretion. Restoration of physiological plasma levels of both estradiol and progesterone returned LH pulse amplitude to values seen in sham OVX controls, and prevented the OVX-induced increase in LH pulse frequency. The group mean LH pulse frequency tended to be less in estradiol + progesterone-treated rats than in sham OVX controls, but this difference was not statistically significant. RU486 blocked uterine progesterone receptors as evidenced by endometrial hemorrhaging. In agreement with the OVX + steroid replacement data, RU486 administration also resulted in increases in LH pulse amplitude and frequency. These data demonstrate that the frequency and amplitude of LH pulses on Day 8 of gestation are held in check by negative feedback signals coming from the ovary. Neither steroid alone exerts any suppressive influence over pulsatile LH secretion during early gestation, but both steroids acting together exert a prominent negative feedback regulation on the pulsatile LH release process.

The object of this study was to examine the influence of both estradiol (E(2)) and progesterone (P) alone or in combination on luteinizing hormone (LH) pulse amplitude and frequency during the interval between Days 21 and 22 of gestation. This was done by analyzing pulsatile LH release in rats bled on Days 21 and 22 of gestation, and in animals ovariectomized (OVX) on Day 21, implanted with silastic capsules producing plasma levels of E(2) and/or P characteristic of the Day 21 to 22 interval, and bled on Day 22 Pulsatile LH release increased between Days 21 and 22 due to an increase in pulse frequency and a small elevation in pulse amplitude. OVX produced no further increase in pulse frequency but markedly enhanced the small change in pulse amplitude. Preventing either the decline in plasma P that normally occurs between Days 21 and 22, or just the small additional decrease in plasma P levels produced by OVX, had no suppressive effect on pulse amplitude or frequency, although Day 22 levels of P alone augmented the normal increase in pulse frequency occurring between Days 21 and 22. Restoration of physiological plasma E(2) levels had no effect on the normal increase in pulse frequency, but partially attenuated the OVX-induced increase in pulse amplitude. Replacement of physiological Day 22 levels of both E(2) and P also decreased LH pulse amplitude, although amplitude was not significantly different from that seen following E(2) replacement alone, and was still greater than the normal Day 22 value. In contrast, restoration of physiological plasma levels of E(2)+ P caused a suppression of LH pulse frequency below that normally seen on Day 22. While E(2)+ P did not completely prevent the OVX-induced increase in pulse amplitude, administration of charcoal-extracted porcine follicular fluid to rats OVX on Day 21, and in which physiological plasma levels of E(2)+ P were restored, caused a further reduction in pulse amplitude. These data demonstrate that 1) marked increases in LH pulse amplitude are prevented from occurring between Days 21 and 22 of gestation by ovarian steroids, notably E(2), and that this suppression is enhanced by a non-steroidal factor present in porcine follicular fluid, 2) neither E(2) or P alone suppresses LH pulse frequency on Day 22 of gestation; LH pulse frequency increases on Day 22 because the plasma level of one of these steroids, P, markedly declines, and 3) restoration of physiological plasma levels of both steroids in the absence of the ovary produces an unphysiological suppression of pulse frequency, i.e. results in a lower pulse frequency than normally occurs in the presence of these same plasma steroid levels in animals with their ovaries intact. One hypothesis consistent with the latter observation is that at the end of gestation in the rat the ovary may produce a factor which 'protects' the frequency of the LH pulse generator from the negative feedback action of ovarian steroids. This allows an increase in LH pulse frequency and mean blood LH levels, and thereby facilitates ovarian follicular development and the normal progress of the first postpartum estrous cycle.

The aim of this study was to examine the regulation of basal pulsatile LH release by ovarian estradiol (E2) and progesterone (P) during the interval between the mornings of proestrus and estrus in the rat estrous cycle. Pulsatile LH release was studied in six groups of rats bled continuously through jugular venous cannulae between 0930-1230 h at a rate of 50 microliter whole blood/5 min: 1) bled on proestrus; 2) sham ovariectomy (OVX) at 0900-1000 h on proestrus and bled on estrus; 3-6) OVX at 0900-1000 h on proestrus, implanted with either empty or E2-, P-, or E2- plus P-containing Silastic capsules, and bled 24 h after OVX. In our colony, plasma E2 levels peaked at 1300 h, remained high through 1730 h, and then declined. Plasma P values increased between 1300 and 1730 h, peaked at 2000 h, and were rapidly declining by 2400 h. To reproduce the magnitude as well as the temporal pattern for these changes in plasma E2 and P levels, E2 capsules were inserted at the time of OVX on proestrus and removed at 1830 h. P capsules were inserted at 1400 h and removed at 2300 h. Groups of ovariectomized or sham-ovariectomized control animals had empty capsules implanted and removed at comparable times. Capsules producing basal E2 and P levels were not inserted after the removal of the original implant, since mean blood LH levels, pulse amplitude, and frequency were the same in rats sham ovariectomized or ovariectomized at 1830 h on proestrus and bled the next morning between 0930-1230 h. Mean blood LH levels decreased between the mornings of proestrus and estrus due to a reduction in LH pulse frequency as pulse amplitude remained stable. OVX at 0900-1000 h on proestrus increased mean blood LH levels 2.5-fold compared to values on estrus due to increases in both LH pulse frequency and amplitude. Restoration of physiological proestrous levels of only E2 returned LH pulse frequency to estrous values, but did not significantly affect LH pulse amplitude. P alone also had no significant effect on LH pulse amplitude, but slightly reduced pulse frequency, although, unlike E2, not to values seen on estrus. Replacing both E2 and P returned LH pulse amplitude to estrous levels and reduced LH pulse frequency.(ABSTRACT TRUNCATED AT 400 WORDS)

Naloxone, an endogenous opioid peptide (EOP) receptor antagonist, increases LH pulse frequency and amplitude in early gestation in the rat (7). The object of this study was to further explore the suppression of pulsatile LH release by EOPs on day 8 of pregnancy by examining whether inhibition of norepinephrine (NE) or epinephrine (EPIN) synthesis, or stimulation of μ-aminobutyric acid (GABA)-B receptors, modified the ability of naloxone infusion (0.5 mg/kg/hr for 3.5 hr) to stimulate pulsatile LH secretion. Blood sampling (50 μl whole blood/5 min) began 0.5 hr after the onset of infusion. Three studies were conducted. 1) LY 134046 (PNMT inhibitor, 50 mg/kg IP), given at − 27, − 20, and − 3 hr relative to the onset of a 3-hr blood sampling period, produced no change in hypothalamic-preoptic area (HPOA) levels of NE, and a 76% decline in HPOA-EPIN levels, as determined by HPLC. Although basal LH pulse frequency was reduced, this treatment had no effect on the stimulatory action of naloxone on pulsatile LH release. 2) FLA-63 (DBH inhibitor, 25 mg/kg IP) given at − 3 hr produced a 72% decline in HPOA-NE levels, a 44% decrease in HPOA-EPIN values, and blocked the stimulatory action of naloxone on LH pulse frequency. Since depletion of HPOA-EPIN by LY 134046 did not compromise the LH response to naloxone, this effect of FLA-63 is due to depletion of HPOA-NE levels. 3) While saline had no effect on the increased pulsatile LH release caused by naloxone, administration of baclofen (GABA-B receptor agonist, 6 mg/kg IV) suppressed the pulsatile LH secretory response to naloxone infusion. These data demonstrate that inhibition of NE but not EPIN synthesis, or stimulation of GABA-B receptors, are both capable of interfering with naloxone-induced stimulation of pulsatile LH release on day 8 of pregnancy in the rat.

The object of this study was to determine the roles of ovarian estradiol (E2) and progesterone (P) in regulating pulsatile LH release between estrus and diestrous day 1 (D1) in the rat estrous cycle. Three groups of rats were bled at rates of 75 or 100 microliter whole blood/6 or 8 min, respectively, between 0930-1230 h on estrus or 24 h after either sham ovariectomy (OVX) on estrus (i.e. on D1) or OVX on estrus. There were no differences in plasma E2 and P levels in rats between estrus and early D1. However, after OVX on estrus, plasma levels of both steroids declined and were significantly lower 24 h later than values in D1 controls, indicating an active ovarian secretion of both hormones in this interval of the rat cycle. A significant increase in mean blood LH levels occurred between estrus and D1 due to an increase in LH pulse amplitude and frequency. After OVX on estrus, all parameters of pulsatile LH release also increased within 24 h, but mean blood LH levels as well as LH pulse amplitude and frequency were virtually identical to values in D1 controls, despite the decline in plasma E2 and P levels. Thus, OVX did not augment the increases in LH pulse amplitude and frequency that occur between estrus and D1. This demonstrates that the increase in pulsatile LH release from estrus to early D1 occurs in the absence of ovarian steroid negative feedback; the increases in LH pulse amplitude and frequency are not under negative feedback control by the low plasma levels of E2 and P present at this time. These data stand in direct contrast to the presence of prominent ovarian steroid negative feedback systems operative between D1 and diestrous day 2, and diestrous day 2 and proestrus.

The aim of this study was to analyze possible estradiol (E 2 )-independent and -dependent endogenous opioid peptide (EOP) suppression of pulsatile LH release between the mornings of diestrus 2 (D2) and proestrus by examining the LH response to naloxone infusions in the presence or absence of proestrous levels of E 2 . Pulsatile LH secretion remained unchanged between D2 and proestrus but mean blood LH levels, pulse amplitude and frequency increased within 24 hr following ovariectomy on D2. This increase was due in large part to the loss of E 2 negative feedback, since restoration of physiological proestrous E 2 levels returned LH pulse frequency to proestrous a.m. levels and greatly reduced pulse amplitude. In ovariectomized rats lacking E 2 negative feedback, continuous infusion of the EOP receptor antagonist naloxone (0.5 and 2 mg/kg/hr) caused a further increase in pulse amplitude and frequency. This naloxone-induced increment in pulsatile LH release was exerted via centrally located EOP receptors since naloxone did not alter pituitary responsiveness to LHRH, and its stimulatory action on pulsatile release was diminished by simultaneous infusion with morphine. Naloxone also increased pulsatile LH release in E 2 -treated animals. The naloxone-induced increments in LH pulse amplitude were the same in the presence or absence of E 2 negative feedback. Moreover, the increments in amplitude produced by naloxone in E 2 -treated rats were significantly less than those resulting from the combination of ovariectomy plus naloxone infusion in empty capsule-implanted rats. These data indicated that naloxone infusion in E 2 -implanted animals blocked an E 2 -independent EOP suppression of this parameter of pulsatile release. In contrast, with both naloxone doses tested, there was a trend for naloxone-induced increments in pulse frequency to be larger in ovariectomized rats previously implanted with E 2 rather than empty capsules. Although these differences were not statistically significant, the naloxone-induced increments in pulse frequency in E 2 -treated rats (unlike those in pulse amplitude) were large enough so that they were also significantly different from the overall increases in pulse frequency caused by the combination of ovarian steroid removal coupled with blockade of EOP receptors during naloxone infusion in empty capsule-implanted animals. This raises the possibility that naloxone infusions in E 2 -treated rats compromised both an E 2 -independent EOP suppression of pulse frequency and a small, E 2 -dependent component as well. Overall, the data demonstrate that between the mornings of D2 and proestrus, EOPs act independent of E 2 to suppress both LH pulse amplitude and frequency, and the negative feedback of E 2 on LH pulse amplitude is not mediated by EOPs whose receptors are antagonized by naloxone. The data also suggest the possibility that E 2 suppression of pulse frequency may be mediated, at least in part, by EOPs.

The objective of this study was to determine if endogenous opioid peptides (EOPs) influence the pattern of pulsatile luteinizing hormone (LH) secretion on days 6-8, 14-16 and 22 of gestation in the rat. Unanesthetized animals with two jugular cannulae were initially infused with 0.9% saline during which the control pattern of pulsatile LH release was determined. Possible EOP involvement was then determined by infusion of the EOP receptor antagonist naloxone. Plasma estradiol (E2) and progesterone (P) values increased between days 6-8 and 14-16. While plasma E2 values remained elevated through day 22, plasma P values declined by 90%. As previously reported, mean blood LH levels during the control period on day 22 were higher than on days 6-8 and 14-16 due to an increase in LH pulse frequency. At each stage of gestation naloxone infusion increased mean blood LH levels. This stimulatory action of naloxone was reduced in a dose-dependent fashion by simultaneous infusion with morphine, demonstrating that this effect is mediated via EOP receptors. There was no difference in the in vivo pituitary responsiveness to LH-releasing hormone (LHRH) between rats infused with saline or naloxone at any stage of pregnancy, demonstrating that the stimulatory effect of naloxone was not exerted at the pituitary level. Naloxone increased both the amplitude and frequency of pulsatile LH secretion on days 6-8, and stimulated frequency on days 14-16. The effect on amplitude could not be assessed on days 14-16 because too few rats exhibited pulsatile LH secretion prior to naloxone infusion. The increase in pulse frequency was similar on days 6-8 and 14-16. Although naloxone increased LH pulse amplitude and frequency on day 22, these increases were significantly less than those seen on days 6-8 and 14-16, respectively. Pituitary responsiveness to LHRH was less at all stages of pregnancy in comparison to responsiveness in ovariectomized rats, and progressively declined from days 6-8 through day 22. The lowest responsiveness to LHRH was seen on day 22 and contributed, at least in part, to the diminished increase in LH pulse amplitude in response to naloxone infusion on day 22 compared to days 6-8. The reduced naloxone-induced increment in LH pulse frequency on day 22, occurring coincident with a precipitous decline in plasma P levels, suggests a decreased EOP suppression of pulse frequency at this time.(ABSTRACT TRUNCATED AT 400 WORDS)

The object of this study was to determine the influence of ovarian steroids on pulsatile LH release in the interval between the mornings of diestrus 2 (D2) and proestrus in the rat. Four groups of rats were bled continuously for 3 h between 09.30–12.30 h at a rate of 75 µl whole blood/6 min: (1) bled on D2; (2) sham ovariectomy (OVX) on D2 and bled on proestrus; (3) OVX on D2, implanted with empty or oil-filled capsules, and bled 24 h later; and (4) OVX on D2, implanted with estradiol (E2) capsules, and bled 24 h later. Between D2 and proestrus, plasma E2 levels increased from 13 ± 1 to 42 ± 9 pg/ml, and progesterone levels decreased from 27 ± 3 to 13 ± 2 ng/ml, the latter reflecting the decline of the corpus luteum early on D2. Between D2 and proestrus there was no change in mean blood LH levels, LH pulse amplitude, or pulse frequency. However OVX on D2 increased mean blood LH levels 2.5-fold over values on proestrus due to a 3.5-fold elevation in LH pulse amplitude and an 80% increase in pulse frequency. E2 levels fell in these rats to 8 ± 1 pg/ml. Restoration of physiological proestrous levels of E2 (46 ± 5 pg/ml) significantly reduced the increase in mean blood LH levels by lowering pulse frequency to proestrous values, and by causing a 50% reduction in pulse amplitude. However, LH pulse amplitude and therefore mean blood LH levels were still higher than values on proestrus. The possibility that ovarian progesterone might also regulate LH pulse amplitude in the D2-proestrous interval was ruled out since there were no significant differences in plasma progesterone levels between 13.00–19.00 h following sham OVX or OVX of rats earlier on the morning of D2. In vitro incubation studies with anterior pituitaries from rats ovariectomized 24 h prior on D2 and implanted with E2 or empty capsules demonstrated that E2 increased the pituitary sensitivity to LHRH, indicating that the E2-induced decrease in LH pulse amplitude could not be exerted at the pituitary level. In conclusion, since LH pulse amplitude and frequency remain unchanged between the mornings of D2 and proestrus, yet increase following OVX, the present data demonstrate that ovarian E2 acts centrally to exert a restraining effect on both parameters of pulsatile LH secretion during this interval in the rat estrous cycle.

Endogenous opioid peptides (EOPs) suppress pulsatile LH release during pregnancy in the rat, but the stimulatory effect of the EOP receptor antagonist naloxone on LH pulse frequency is reduced or eliminated on day 22 of gestation. Plasma progesterone (P) levels are elevated through day 20 and fall by day 22. The aim of this study was to determine whether the decline in plasma P levels underlies the loss of EOP suppression of LH pulse frequency on day 22. Rats were bled on day 20 of pregnancy while being infused with 0.9% saline (0.5 ml/h) for 3 h, or implanted with empty or P-filled silastic capsules on day 20 and bled on day 22 while being infused first with saline for 3 h and then naloxone (0.5 mg/kg/h) for 3 h. Plasma P levels in the P-capsule group did not differ significantly from day 20 values, whereas P values in the empty capsule group were markedly decreased compared to day 20 levels and to values in the P-capsule group. Plasma estradiol values did not vary significantly between the two capsule-implanted groups. Mean blood LH levels increased between day 20 and day 22 due to an increase in LH pulse frequency and a small but significant increase in LH pulse amplitude. On day 22, mean blood LH levels, pulse amplitude and pulse frequency values during the saline infusion period in the P-capsule group were less than in the empty capsule group, and did not differ from values in the day 20 group.(ABSTRACT TRUNCATED AT 250 WORDS)

Highly purified gonadotropin preparations from representatives of all four tetrapod classes and several fish were used to examine the specificity of in vitro testosterone production by minced testes from four species of teleost fish, representing three families and two orders. No consistent phylogenetic patterns emerged in either hormonal (FSHLH) or phylogenetic specificity in the steroidogenic response of the teleost testis. Testes of Gillichthys mirabilis (Family Gobiidae) were highly specific for all species of LH (FSHs were inactive) and they also showed a certain degree of species specificity; mammalian and amphibian LHs were much more potent than reptilian and avian LHs. In contrast, testes of Cichlasoma citrinellum and Sarotherodon mossambicus (Family Cichlidae) lacked gonadotropin specificity (most preparations of FSH and LH were about equipotent), and in C. citrinellum all species of tetrapod hormones were active within approximately the same dose range. Gonadotropins from the sturgeon (Chondrostei) were the most potent in both G. mirabilis and C. citrinellum. In Salmo gairdneri (Family Salmonidae) there was specificity for amphibian (bullforg) LH, but not for mammalian (ovine) LH. Steroid secretion by the fish testes was temperature dependent, but relative potencies of ovine LH, bullfrog LH, and sturgeon gonadotropin were not altered by incubation temperatures between 10 and 30° in G. mirabilis. As a group, the teleosts examined here differ from mammals, birds, and reptiles in their hormonal specificity (or consistent lack there of) for testicular androgen secretion; they most closely resemble the Amphibia in their high degree of interspecific variability in response to FSH and LH. The lack of predictability in this aspect of specificity as well as in species specificity in the potencies of individual types of hormones of different origins precludes generalizations about the evolutionary pattern of gonadotropin-testis interactions.

Clearance of exogenous gonadotropins was studied in the lizard Anolis carolinensis in relation to the problem of variations in potency ratios between LH and FSH of different species of hormone. Studies with unlabeled bullfrog ( Rana catesbeiana ) gonadotropins revealed that the FSH had a much longer persistence time in the lizard than did LH; this was confirmed by direct immunological measurements of circulating hormone and by temporal profiles in the stimulation of gonadal androgen production in the lizard. Bullfrog FSH and LH labeled with 125 I had similar clearance rates; the clearance of ( 125 I) Rana LH was slower than that of the unlabeled preparation. In contrast, similar studies with unlabeled sea turtle ( Chelonia mydas ) hormones indicated that the LH is cleared more slowly than is FSH. These differences between the pairs of frog and turtle gonadotropins are consistent with the difference in LH/FSH potency ratios observed for the two species of hormone in the in vivo Anolis lizard testes weight bioassay. Thus, these data provide additional insights into the variability in effects of different species of gonadotropins: the relatively high potency of some species of LH may be related in part to increased half-lives.

Competitive binding assays using 125I-labeled equine LH confirm that this hormone binds with relatively high affinity to both LH- and FSH-binding sites on the separated Leydig cells and seminiferous tubules, respectively, of the 18-day-old rat testis. In contrast, ovine LH is potent only in Leydig cell LH-binding sites. However, despite its binding to FSH sites, equine LH, like all other LH preparations examined, lacks the ability to stimulate in vitro cAMP production from the rat seminiferous tubules, a characteristic action of FSH. Although all three species of LH examined (equine, ovine, and human) are similarly inactive in stimulating cAMP production when tested alone (all were

SUMMARY The actions of human follicle-stimulating hormone (hFSH) and its β-subunit were examined in several assays in reptiles, including effects on lizard testicular activity (growth and androgen production) in vivo, and stimulation of androgen production by snake testes and competition for binding of 125I-labelled hFSH in lizards and snakes in vitro. Binding was also examined with mammalian tissues. The hFSH was highly steroidogenic in the snake and lizard; otherwise results were similar to those observed in mammals. In all cases, the potency of the β-subunit was only a few per cent of the intact hormone. The potency of hFSH in vivo compared with NIH-FSH ovine standards was several 100 times greater than in vitro. Results for stimulation of androgen production in vivo closely paralleled those for binding assays in both reptiles and mammals. In contrast to previous results for ovine FSH β-subunit, human FSH β-subunit has little if any FSH biological activity in reptiles.

Fractionation of pituitary glands of the ostrich (Struthio camelus) resulted in the preparation of highly purified growth hormone (GH), prolactin (PRL), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) in relatively high yields; thyrotropin (TSH) was partially purified. Hormones were identified by a variety of biological and immunological tests. Biochemical and immunological characterizations were performed to study the purity of each preparation and to allow comparison with other species of hormones, with special emphasis on other avian hormones. Similarities to the corresponding hormones of other tetrapods and even fish are apparent in the various biochemical features. THe ostrich hormones show closer affinities to other avian hormones than to mammalian or reptilian species in some respects (especially immunological), but notable biochemical differences are also evident among the few avian species examined.

An antiserum generated against ovine follicle-stimulating hormone (FSH) was used to examine phylogenetic relatedness of FSH molecules among the four classes of tetrapods. Heterologous radioimmunoassay (RIA) employing this antiserum with 125I-labeled human FSH as tracer revealed a high degree of immunochemical similarity among purified FSH molecules from diverse eutherian, metatherian, avian, and reptilian species. While the eutherian hormones tended to be slightly more potent than those from other amniotes, at least one avian FSH (ostrich) exhibited an equivalent degree of antigenicity. Hormones from amphibians formed a discrete group with lower cross-reactivity. In all of these species, only the gonadotropin previously identified as an FSH on the basis of physicochemical and biological profiles showed appreciable cross-reactivity; LH preparations were essentially inactive. 125I-Labeled FSH from a bird (turkey), reptile (sea turtle), and amphibian (bullfrog) bound specifically to the anti-ovine FSH serum, and RIAs with these as tracers yielded essentially the same results as outlined above. Immunoneutralization tests employing binding of 125I-labeled hormone to gonadal receptors confirmed that the anti-ovine FSH serum cross-reacted with the biologically active form of the FSH from the human, bird, and turtle. Moreover, the antiserum appeared to preferentially bind to that portion of the radiolabeled hormone that exhibited the greatest capacity to bind to tissues. The squamate reptiles (snakes and lizards) presented a striking exception to the above phylogenetic patterns, since no antigenic cross-reactivity could be detected with the pituitaries or purified gonadotropins from these reptiles.

Biological and binding activities of adenohypophysial hormones purified from the ostrich (ost), Struthio camelus, were compared to those of the corresponding hormones derived from mammalian and other avian species. The potency of ostrich prolactin was comparable to those of other avian preparations and slightly less active than the ovine hormone when tested in the pigeon crop-sac assay, but ostrich growth hormone (GH) was more potent than several other avian preparations and was comparable to mammalian GH in the rat tibia bioassays. Marked discrepancies were evident in the activities of both ostrich gonadotropins (ostGn) when they were tested in a variety of in vivo and in vitro bioassays and radioreceptor assays (RRAs). Both the ostrich follicle-stimulating hormone (ostFSH) and luteinizing hormone (ostLH) were among the most potent tested thus far in in vivo bioassays for total gonadotropin in a lizard and a cockerel; a high sialic acid content may account for these high potencies. OstFSH was also the most potent nonmammalian Gn tested in two FSH specific mammalian bioassays, an in vivo (ovarian augmentation) and an in vitro (cAMP production) rat bioassay; in fact, ostFSH behaved more like a mammalian than an avian hormone in these assays. However, the binding activity of ostFSH was not unlike that of other avian FSH preparations when tested in either mammalian or nonmammalian FSH-RRA systems. OstLH was more potent than other avian preparations in an in vitro mammalian bioassay, but not in avian or amphibian LH bioassays; species specificity was pronounced among these LH assays. Binding activities of ostLH in mammalian and avian LH-RRAs were generally consistent with potencies in the two species of bioassays. However, a marked discrepancy was apparent in the behavior of ostLH in FSH-RRAs. Although ostLH had very low FSH activity when tested by radioimmunoassay, by bioassay, or by FSH-RRA with avian gonads, it was equipotent to ostFSH in competing for FSH-binding sites on the mammalian gonad; in this respect it was more like turkey than chicken LH. The ability of ostLH to antagonize the biological activities of ostFSH in the stimulation of cAMP by rat seminiferous tubules confirms that ostLH binds to the same functional receptors as FSH on the rat testis, even though it does not induce the characteristic physiological response associated with FSH.

Gonadotropin preparations from three classes of tetrapods (amphibian, avian, and mammalian) and a chondrosteian and teleost fish were used to investigate the species specificity and hormonal ( FSH LH ) specificity of in vitro steroid (estradiol-17β) production by the teleost ovary. Results for ovaries from one species of gobiid, Gillichthys mirabilis , and two cichlids, Cichlasoma citrinellum and Sarotherodon mossambicus , revealed a general lack of species specificity in the response to tetrapod gonadotropins, but the piscine, especially sturgeon, gonadotropins were much more potent than any of the tetrapod hormones. All three species of teleost ovaries responded to both types of tetrapod gonadotropins (FSH and LH), but the extent of hormonal specificity was variable. The gobiid ovary showed the highest LH specificity (potencies of FSHs = 7–14% of LH); in the two cichlids the potency of FSHs ranged from 11 to 100% of the respective LHs. In general, the specificity of the ovarian steroidogenic response to gonadotropins parallels that observed for testosterone secretion in the males of the same three fish, but the differential actions of the tetrapod hormones (both species and hormonal specificity) are more exaggerated for the testes.

Follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were purified from the pituitary glands of the turkey (Meleagris gallopavo). These hormones were characterized biochemically and biologically and compared with chicken gonadotropins prepared in several independent laboratories. Amino acid and carbohydrate analyses demonstrated homology between turkey and other species of gonadotropin. The turkey LH purified here had significantly higher carbohydrate content than a previous preparation of turkey LH. Immunological studies further confirmed that the turkey FSH and LH were distinct from one another and that each was homologous to the respective gonadotropin from other vertebrates; the immunopotencies of the turkey hormones were similar to those from chickens. A variety of bioassays and radioreceptor assays (RRAs) confirmed the biological activity of the two turkey gonadotropins and revealed that the turkey LH was distinct from that of the chicken. As expected, the two types of turkey hormones were approximately equipotent in total gonadotropin bioassays (frog spermiation and 32P uptake by chick testes), and only the turkey LH was active in the rat Leydig cell assay and in RRA for LH in mammals. However, the turkey LH was also highly potent in several assays considered to be relatively FSH specific, including the Anolis lizard assay and several RRA systems using mammalian, turtle and avian gonadal receptors with 125I-labeled human FSH as tracer. Turkey and chicken FSH are similar in the RRAs, but the turkey LH was consistently more potent than either avian FSH in competing for FSH-binding sites. Chicken LH had relatively low activity by comparison. It is suggested that the evolution of the structure of active sites in turkey LH has involved convergence on those of the FSH molecule.

This study examined the importance of pulsatile luteinizing hormone (LH) release on diestrus 1 (D1; metestrus) in the rat estrous cycle to ovarian follicular development and estradiol (E2) secretion. Single injections of a luteinizing hormone-releasing hormone (LHRH) antagonist given at -7.5 h prior to the onset of a 3-h blood sampling period on D1 reduced mean blood LH levels by decreasing LH pulse amplitude, while frequency was not altered. Sequential injections at -7.5 and -3.5 h completely eliminated pulsatile LH secretion. Neither treatment altered the total number of follicles/ovary greater than 150 mu in diameter, the number of follicles in any size group between 150 and 551 mu, or plasma E2, progesterone, or follicle-stimulating hormone (FSH) levels. However, both treatments with LHRH antagonist significantly increased the percentage of atretic follicles in the ovary. These data indicate that: 1) pulsatile LH release is an important factor in determining the rate at which follicles undergo atresia on D1; 2) reductions in LH pulse amplitude alone are sufficient to increase the rate of follicular atresia on D1; 3) an absence of pulsatile LH release for a period of up to 10 h on D1 is not sufficient to produce a decline in ovarian E2 secretion, most likely because the atretic process was in its early stages and had not yet affected a sufficient number of E2-secreting granulosa cells to reduce the follicle's capacity to secrete E2; and 4) suppression or elimination of pulsatile LH release on D1 is not associated with diminished FSH secretion.

The object of this study was to further define the roles of both norepinephrine (NE) and epinephrine (EPIN) in regulating pulsatile luteinizing hormone (LH) release in 4-day ovariectomized rats, in particular to examine the effect of decreasing NE synthesis on pulsatile LH secretion in animals with already greatly depleted levels of brain EPIN. Rats were injected ip with vehicle or drug at -27, -20, -5 and - 3 h relative to the onset of a 3-h blood sampling period. Hypothalamic-preoptic area (HPOA) levels of NE and EPIN were determined by high-performance liquid chromatography. Compared to controls, FLA-63 (25 mg/kg, a dopamine-ss- hydroxylase inhibitor), given at - 3 h, produced 50% and 22% declines in HPOA-NE and EPIN, respectively, and reductions in pulse amplitude and frequency. LY134046 (50 mg/kg, a phenylethanolamine N-methyltransferase inhibitor), given at - 27, - 20 and - 5 h, or -27, -20, -5 and -3 h, produced no change in NE, 88% and 86% declines in EPIN, respectively, and reductions in pulse frequency only. Each LY134046 treatment protocol produced the same decline in EPIN and pulse frequency. Thus, EPIN levels were maximally decreased by three LY134046 injections. When rats were given LY134046 at -27, -20 and -5 h, and FLA-63 at -3 h, compared to rats treated with LY134046 alone, there was no further decrease in HPOA-EPIN (82% decline), a 46% decline in NE, a further reduction in pulse frequency and a reduction in pulse amplitude. This further suppression of LH release must be due to a reduction in HPOA-NE levels since no further decrease in EPIN levels occurred. These data demonstrate within the same animal that NE and EPIN are both stimulatory to pulsatile LH release. NE stimulates the amplitude and frequency, and EPIN stimulates the frequency of pulsatile LH secretion.

The object of this study was to examine the effect of porcine follicular fluid (PFF) alone or in combination with estradiol (E2) on pulsatile LH release during the interval between the mornings of diestrus 2 (D2) and proestrus in the rat. Steroids were removed from PFF by charcoal extraction. Preliminary studies indicated that 1 ml PFF given intraperitoneally suppressed FSH secretion for up to 15 h, with an onset of action between 3 and 4 h and maximal suppression between 6 and 9 h. In subsequent experiments, six groups of animals were bled continuously for 3 h between 07.30 and 10.30 h at a rate of 50 microliter whole blood/5 min: group 1 was bled on D2; group 2 was sham ovariectomized on D2 (08.30-09.30 h), immediately implanted with an empty capsule, given saline at 12.00 and 24.00 h, and bled on proestrous AM; groups 3-6 were ovariectomized on D2, implanted with an empty or E2 capsule, given 1 ml saline or PFF at 12.00 and 24.00 h, and bled 24 h following ovariectomy (OVX). Between D2 and proestrus plasma E2 levels increased, and there was no change in any parameter of pulsatile LH release. However, OVX on D2 reduced plasma E2 levels and increased mean blood LH levels above proestrous values due to increases in LH pulse amplitude and frequency. Restoration of physiological proestrous levels of E2 reduced the increase in mean blood LH levels, by lowering pulse frequency to proestrous values and by greatly reducing pulse amplitude. However, LH pulse amplitude and mean blood LH levels were still higher than values on proestrus. PFF alone produced no alteration in any parameter of pulsatile LH release compared with saline-treated animals.(ABSTRACT TRUNCATED AT 250 WORDS)

The anterior pituitary hormones LH, GH, and PRL were purified from several marsupial species, Eastern grey kangaroo ( Macropus giganteus ), Western grey kangaroo ( Macropus fuliginosus ), tammar wallaby ( Macropus eugenii ), and brush possum ( Trichosurus vulpecula ). LH was isolated only from the kangaroos, whereas GH and PRL were isolated from all four species. The purified marsupial hormones were identified on the basis of bioassay, radioreceptor assay, and radioimmunoassay. They were found to be less potent biologically than the respective eutherian hormones in eutherian assays. Chemical characterization, including NH 2 -terminal analysis, amino acid composition, disc gel electrophoresis, and molecular weight determination, revealed basic similarities to eutherian hormones. A homologous, double-antibody radioimmunoassay for Eastern grey kangaroo PRL was developed. This assay was specific for marsupial PRLs; there was very low or no cross-reaction with other kangaroo hormones or eutherian PRLs. In general, the marsupial hormones were found to resemble their eutherian counterparts, but there were differences in biological and immunological potencies, emphasizing the need for purified marsupial hormone standards in studies of marsupial endocrine systems.

The object of this study was to determine whether the increase in LH pulse frequency and mean blood LH levels on day 22 of pregnancy in the rat is due to the precipitous fall in plasma progesterone (P) levels that occurs late in gestation. On day 20 of pregnancy two groups of animals with indwelling jugular cannulae were implanted sc with empty or P-containing Silastic capsules. Blood samples were withdrawn 0.5 h before and 5.5 h postimplantation on day 20 (0800 and 1400 h), at 1400 h on day 21, and at the end of the study between 1200-1300 h on day 22 to follow the time course of changes in plasma P levels over this 2-day period in both groups. These groups were bled on day 22 for 3 h between 0900-1200 h for analysis of pulsatile LH release. A third group not implanted with Silastic capsules was bled on day 20 for 3 h; plasma P levels in these rats bled on day 20 did not differ from the preimplantation values observed in either group of capsule-implanted rats. In empty capsule-implanted animals, plasma P values declined slightly from days 20 to 21 and were dramatically reduced between days 21 and 22. In contrast, after implantation of P capsules, plasma P levels were elevated on day 20 and remained elevated on day 21 compared with preimplantation values. Although these increased plasma P values declined between days 21 and 22, reflecting a decrease in endogenous P secretion, they were nonetheless comparable to day 20 values due to the presence of the P-containing capsules. Plasma estradiol values did not differ significantly between any of the experimental groups. In the empty capsule group bled on day 22, mean blood LH levels and LH pulse frequency were significantly higher compared to day 20 values, at a time when plasma P levels had fallen significantly from day 20 values. However, in the P capsule group, mean blood LH levels and LH pulse frequency on day 22 were significantly lower than values in the empty capsule group and were not different from the low values on day 20. Thus, preventing a decline in plasma P values to the low levels normally found on day 22 prevented the increase in LH pulse frequency and mean blood LH levels normally seen at this time of pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)

Conventional fractionation procedures used for separation and purification of pituitary FSH and LH in diverse tetrapods were employed to study pituitary gonadotropin (Gn) from five genera of snakes representing three families: Elaphe and Ptyas—Colubridae; Naja and Bungarus—Elapidae; Crotalus—Viperidae. The hormones purified from each of these snakes were shown to be relatively potent in a variety of Gn bioassays and radioreceptor assays (RRAs) in snakes but there was no clear evidence for two separate types of Gn molecule. These snake materials were also active in RRAs and bioassays (testis growth, androgen production, and ovulation) in lizards, but poor dose-response characteristics and relatively low potencies were observed in most cases. An even higher degree of species specificity was evident when snake hormones were tested in nonsquamate species, including other reptiles. Most snake Gn fractions were essentially inactive in all RRAs and bioassays employing amphibians, birds, mammals, and turtles; these assays included several that typically show broad species cross-reactivity (e.g., anuran ovulation and spermiation, 32P uptake by chick testis, and in vitro androgen production in birds and turtles). Radiolabeled Gn from a snake (Naja) bound specifically only to gonadal receptors from snakes and lizards. Biochemical analyses of the snake Gn confirmed their glycoprotein nature but failed to show clear structural homology to either FSH or LH. Chromatographically, the purified Gn from Naja naja tended to behave predominantly like an LH and its electrophoretic mobility on polyacrylamide gels also suggested an LH-like molecule, but several problems complicated interpretation of these results. Amino acid composition of this snake Gn revealed similarities to both FSH and LH, but it was not consistently like either. Radioimmunological studies (RIAs) with several heterologous gonadotropin antisera also failed to show a consistent relatedness between snake hormones and either FSH or LH; in fact, the snake Gns are exceptional among tetrapods in showing a lack of cross-reactivity in several FSH and LH-RIA systems. Antisera were raised against Gn from two species of snake. Antiserum to Naja Gn blocked the biological and binding activities of other species of snake Gn and it also selectively neutralized the activity of FSH (but not LH) from a turtle, alligator, and bird. However, in homologous RIA, the Naja Gn showed a slight cross-reaction only with another elapid species. In a heterologous RIA (using antiserum to Ptyas Gn and 125I-labeled Naja Gn), relatively high cross-reactivity was seen with Gn from several elapids and colubrids but not viperids or any non-snake species. Thus, the snake hormones show partial relatedness to one another but are generally distinct from FSH or LH; the only evidence of cross-reaction with heterologous (non-serpentine) gonadotropin suggests that they share common immunochemical determinants with reptilian FSH. Overall, the snake gonadotropins appear to be unique among tetrapod gonadotropins in terms of their biological cross-reactivity, biochemical composition, and immunochemical properties. Data suggest that snakes may only have a single gonadotropin that does not show a clear homology to either FSH or LH.

We have previously demonstrated an absence of ovarian steroid negative feedback on pulsatile luteinizing hormone (LH) release between estrus and early diestrus 1 (D1) in the rat estrous cycle. The object of the present study was to determine if there was a steroid-independent endogenous opioid peptide (EOP) suppression of pulsatile LH release in thes same 24-h interval, and if so, which parameter(s) of pulsatile LH release were affected. Rats were bled on estrus, or 24 h following sham ovariectomy (OVX), or OVX at 08.30–10.00 h on estrus. At the time of bleeding all rats were infused i.v. for 4 h either with 0.9% saline (0.5 ml/h) or naloxone (0.005, 0.05, 0.5, or 2 mg/kg/h). At 1 h after the infusion began, rats were bled for 3 h (40 or 50 μl whole blood/5 min) between 09.30 and 12.30 h. Mean blood LH levels increased between estrus and early D1 due to increases in LH pulse amplitude and frequency. OVX on estrus decreased plasma levels of estradiol and progesterone 24 h later, but did not augment the increase in pulsatile LH release. However, naloxone infusion augmented the increase in pulsatile LH secretion in sham ovariectomized rats in a dose-dependent fashion. While infusion of 0.005 or 0.05 mg/kg/h had no effect, 0.5 or 2 mg/kg/h increased blood LH levels by increasing both LH pulse amplitude and frequency. The stimulatory effect of naloxone on pulsatile LH release was blocked by simultaneous infusion of morphine, demonstrating that the effect was mediated by EOP receptors. In addition, pituitary responsiveness to LHRH in vivo was not altered by naloxone infusion, indicating a central site of action for the EOP receptor antagonist. These data confirm our previous report that the increases in LH pulse amplitude and frequency occuring from estrus to early D1 occur in the absence of ovarian steroid negative feedback. However, pulsatile LH release would increase even further in this time interval were it not for the existence of a centrally occurring, steroid-independent EOP suppression of both LH pulse amplitude and frequency during this 24-h period of the rat estrous cycle.

This study examined the importance of pulsatile luteinizing hormone (LH) release on diestrus 1 (D1; metestrus) in the rat estrous cycle to ovarian follicular development and estradiol (E2) secretion. Single injections of a luteinizing hormone-releasing hormone (LHRH) antagonist given at -7.5 h prior to the onset of a 3-h blood sampling period on D1 reduced mean blood LH levels by decreasing LH pulse amplitude, while frequency was not altered. Sequential injections at -7.5 and -3.5 h completely eliminated pulsatile LH secretion. Neither treatment altered the total number of follicles/ovary greater than 150 mu in diameter, the number of follicles in any size group between 150 and 551 mu, or plasma E2, progesterone, or follicle-stimulating hormone (FSH) levels. However, both treatments with LHRH antagonist significantly increased the percentage of atretic follicles in the ovary. These data indicate that: 1) pulsatile LH release is an important factor in determining the rate at which follicles undergo atresia on D1; 2) reductions in LH pulse amplitude alone are sufficient to increase the rate of follicular atresia on D1; 3) an absence of pulsatile LH release for a period of up to 10 h on D1 is not sufficient to produce a decline in ovarian E2 secretion, most likely because the atretic process was in its early stages and had not yet affected a sufficient number of E2-secreting granulosa cells to reduce the follicle's capacity to secrete E2; and 4) suppression or elimination of pulsatile LH release on D1 is not associated with diminished FSH secretion.

Torpor was monitored daily in adult male and female European hamsters (Cricetus cricetus) induced to hibernate by exposure to a cold environment (6 degrees C). The effect of photoperiodic manipulations or administration of exogenous gonadal steroids was examined in gonadectomized or intact hamsters. 1. Gonadal regression occurred in all short day, but only in some long day, cold-exposed hamsters. Entry into hibernation was not observed until reproductive regression had occurred. Thus, gonadal atrophy appears to be a necessary precondition for hibernation. 2. Castrated hamsters in the short day cold condition showed a significantly greater incidence of torpor than those in the long day cold condition. Hence, photoperiod affected torpor independently of its effect on the gonadal cycle. 3. Testosterone, when administered via silastic capsules at near physiological levels, completely inhibited torpor in gonadectomized male and female hamsters hibernating in the short day cold condition. 4. In ovariectomized females, torpor was unaffected by progesterone treatment, but partially inhibited by estradiol. A greater inhibition of torpor was observed when estradiol-primed females were administered both estradiol and progesterone simultaneously. Thus, the effect of both hormones may be functionally comparable to that of the single testicular hormone. 5. Estradiol inhibited torpor to a greater extent in intact and ovariectomized female hamsters hibernating in long days than those in short days, suggesting an effect of photoperiod on responsiveness to estradiol. These results indicate an inverse relationship between the gonadal and hibernation cycles, and a probable role for gonadal steroids to influence the timing of the hibernation season. However, non-gonadal factors must also be involved in controlling hibernation, since photoperiod affected the incidence of torpor in gonadectomized animals and because hamsters were able to terminate hibernation in the absence of gonadal hormones.

Follicle-stimulating hormone (FSH) from the ostrich, Struthio camelus, was dissociated by methods previously used to prepare subunits from mammalian gonadotropins. Two chemically dissimilar subunits were obtained from the ostrich FSH and these resembled the alpha- and beta-subunits of mammalian FSH in amino acid composition. The subunits were relatively inactive in radioimmunoassay, radioreceptorassay, and bioassay when tested alone, but significant activity was regenerated upon their recombination. Incubations of mixtures of one of the ostrich subunits with the opposing subunit of ovine FSH also resulted in a significant regeneration of binding and biological activity in FSH assays. These hybrid recombinants demonstrated that the species specificity of the FSH molecule is conferred entirely by the source of the beta-subunit; in fact, hybrids formed between ovine LH-alpha and either ovine or ostrich FSH-beta were more potent in FSH assays than were those containing even two homologous FSH subunits. In contrast, there was little regeneration of LH bioactivity when FSH subunits (from ostrich or sheep) were combined with the subunits of ovine LH, although some LH binding activity was obtained when the mixture contained either one of the LH subunits; i.e., even LH-alpha + FSH-beta showed significant enhancement of binding activity for LH receptors. Thus, the subunits of avian FSH show biochemical and functional homologies to those of mammalian FSH, but there is only limited interchangeability with ovine LH subunits.

Subunits of turkey luteinizing hormone (LH) were prepared from highly purified turkey LH by countercurrent distribution (CCD) and Sephadex G-100 chromatography. The material with the low partition coefficient was designated turkey LH-α and that with the high partition coefficient was designated turkey LH-β. The LH-α eluted from Sephadex G-100 with a Ve Vo ratio of 2.41, while the LH-β eluted at 2.33. The α-subunit had more lysine than arginine, the reverse is seen in the LH-β. The β-subunit has a higher content of leucine, proline, valine, and glycine than the α, but appears to contain no methionine and probably no histidine. Both subunits are high in half-cystine residues. The bulk of the carbohydrate was associated with LH-α (14.9%), while the LH-β had a low carbohydrate content (4.4%). The isolated subunits had very low immuno- and bioactivity, but activity in radioimmunoand radioreceptor assays was regenerated by coincubation of the turkey subunits. Hybridization of turkey LH subunits with those from ovine LH also generated radioreceptor activity. The hybrid prepared with ovine LH-α and turkey LH-β was quite active in an avian testes radioreceptor assay; the reciprocal recombinant was not. The recombinant formed by turkey LH-α and ovine LH-β was active in a mammalian radioreceptor assay. The recombinant formed from turkey LH-α and ovine LH-β was quite active in a Leydig cell testosterone production assay, the reciprocal recombinant was not. Turkey LH is thus shown to be made up of two dissimilar subunits, each of which has substantial chemical homology with those from ovine LH. The species specificity of these two tetropod luteinizing hormones appears to be associated with the β-subunit.

A glycoprotein of neurohypophysial origin was found to have cofractionated with FSH prepared from pituitary glands of the green turtle, Chelonia mydas. Antiserum raised against this preparation contained high antibody titres and affinity for the neurohypophysial component and allowed development of a specific radioimmunoassay to monitor its purification and distribution in the brain. Immunocytochemistry revealed that the glycoprotein was concentrated in the pars nervosa and associated nerve tracts passing through the median eminence to the supraoptic and paraventricular nuclei; similar distributions were observed in turtles and rats. The antiserum to the turtle material bound radiolabelled rat vasopressin (VP)-neurophysin and precipitated precursors of this neurophysin, but it did not cross-react with rat oxytocin-neurophysin. An amino-terminal alanine was also consistent with the structure of rat VP-neurophysin, but the turtle molecule was larger than the corresponding rat molecule. Limited tryptic digests of the turtle glycoprotein contained two components, one of which bound to lysine VP. Both components contained carbohydrate, but only the one which bound to VP cross-reacted in a radioimmunoassay for rat VP-neurophysin. The apparent surge in plasma immuno-FSH at the time of oviposition previously described in the turtle probably represented release of a neurophysin-like 'carrier' molecule associated with secretion of the neurohypophysial hormone (e.g. arginine vasotocin; AVT) responsible for oviduct contractility. These data suggest that the neurohypophysial glycoprotein represents a partially processed AVT precursor and provide the first biochemical evidence of a mammalian-like biosynthetic pathway for neurohypophysial hormones in a non-mammalian species. J. Endocr. (1984) 103, 97–106

We have previously demonstrated an absence of ovarian steroid negative feedback on pulsatile luteinizing hormone (LH) release between estrus and early diestrus 1 (D1) in the rat estrous cycle. The object of the present study was to determine if there was a steroid-independent endogenous opioid peptide (EOP) suppression of pulsatile LH release in this same 24-h interval, and if so, which parameter(s) of pulsatile LH release were affected. Rats were bled on estrus, or 24 h following sham ovariectomy (OVX), or OVX at 08.30-10.00 h on estrus. At the time of bleeding all rats were infused i.v. for 4 h either with 0.9% saline (0.5 ml/h) or naloxone (0.005, 0.05, 0.5, or 2 mg/kg/h). At 1 h after the infusion began, rats were bled for 3 h (40 or 50 microliters whole blood/5 min) between 09.30 and 12.30 h. Mean blood LH levels increased between estrus and early D1 due to increases in LH pulse amplitude and frequency. OVX on estrus decreased plasma levels of estradiol and progesterone 24 h later, but did not augment the increase in pulsatile LH release. However, naloxone infusion augmented the increase in pulsatile LH secretion in sham ovariectomized rats in a dose-dependent fashion. While infusion of 0.005 or 0.05 mg/kg/h had no effect, 0.5 or 2 mg/kg/h increased blood LH levels by increasing both LH pulse amplitude and frequency. The stimulatory effect of naloxone on pulsatile LH release was blocked by simultaneous infusion of morphine, demonstrating that the effect was mediated by EOP receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

The present studies were designed to characterize LH release during pregnancy in the rat. Unanesthetized animals with jugular cannulae were bled for 3 h between 1000-1300 h on days 6-8, 14-16 or 22 of gestation (50 microliters whole blood/5 min). Plasma estradiol and progesterone values both increased from days 6-8 to days 14-16. However, while plasma estradiol levels increased further between days 14-16 and day 22, plasma P levels had declined 86%. The percent coefficients of variation obtained for alterations in blood LH levels at each stage of pregnancy were all significantly greater than intraassay variation, indicating that LH release was pulsatile at each stage. Although there were no significant differences in mean blood LH levels, pulse amplitude, or frequency between days 6-8 and 14-16, the individual patterns of LH release clearly varied between these 2 groups, and most notably within the 14-16 day group. Fifty-three percent (9 of 17) of the LH records in rats on days 14-16 were nonpulsatile compared to only 20% (3 of 15) on days 6-8. However, despite a trend toward an absence of pulsatile LH release on days 14-16, mean frequency at this time did not differ from days 6-8, since on days 14-16 the remaining 8 animals demonstrated 3.5 pulses/3 h, while on days 6-8 the other 12 rats averaged only 2.5 pulses/3 h. On day 22, there was a marked increase in mean blood LH levels compared with either days 6-8 or 14-16. This increase was due to an increase in mean LH pulse frequency. All 15 rats demonstrated pulsatile LH secretion, a significantly greater incidence of pulsatile LH release than on days 14-16 (100% vs. 47%). These data demonstrate that LH release is pulsatile during pregnancy in the rat, and changes in the characteristics of this secretion occur at different stages of gestation.

The aim of this study was to determine whether the rapid increases in LH pulse amplitude and frequency that occur within 24 h after ovariectomy (ovx) on diestrus day 1 (D1) were due to the removal of progesterone (P) and/or estradiol (E). Initial studies demonstrated that plasma levels of E and P were 18.2 +/- 1.2 pg/ml and 34.1 +/- 3.2 ng/ml, respectively, between the evening of D1 and the morning of D2 in our colony of intact rats. Immediately after ovx and jugular venous cannulation on the morning of D1, rats were implanted either with empty Silastic capsules or capsules capable of restoring physiological levels of E and P to the control values reported above. These rats were continuously bled (75 microliter/6 min) for 3 h 1 day after ovx for analysis of pulsatile LH release, and then additional plasma samples were gathered for determination of E and P levels. Rats with empty capsules had decreased levels of E and P and increases in mean blood LH levels, LH pulse amplitude, and pulse frequency. Animals with E capsules had physiological levels of E and decreased levels of P, but no suppression of the acute post-ovx increase in pulsatile LH release. In contrast, animals with P capsules had physiological plasma levels of P, decreased levels of E, and a marked reduction in the acute LH response to ovx. This suppression was due entirely to a decrease in LH pulse amplitude, as pulse frequency was not altered. Rats with E and P capsules had physiological levels of these hormones, which resulted in an even greater reduction in the acute LH response to ovx. This suppression was due to decreases in both LH pulse amplitude and pulse frequency. The effect of P on LH pulse amplitude was centrally mediated, since the in vitro response to LHRH of anterior pituitary fragments from P-implanted rats was the same as that of anterior pituitary fragments taken from rats with empty capsules. These studies demonstrate that the acute increase in LH pulse amplitude that occurs within 24 h after ovx on D1 is due to the absence of a central inhibitory effect of ovarian P, while the rapid increase in LH pulse frequency is due to the loss of both ovarian E and P.(ABSTRACT TRUNCATED AT 400 WORDS)