Gonadotroph-lactotroph associations and expression of prolactin receptors in the equine pituitary gland throughout the seasonal reproductive cycle

An interaction between gonadotroph and lactotroph cells of the pituitary gland has long been recognized in several species. The current study was conducted to investigate whether an association between gonadotrophs and lactotrophs occurs in mares and whether prolactin receptors are expressed within the pituitary gland of this species. The effects of both reproductive state and season on these variables were examined in pituitary glands obtained from sexually active mares in July (breeding season), sexually active mares in November (non-breeding season) and anoestrous mares in November. Pituitaries were dissected out immediately after death and immunofluorescent staining was carried out on 6 micrometer sections using specific antibodies to the LHbeta subunit, FSHbeta subunit, prolactin and prolactin receptor. Gonadotrophs were observed in both the pars distalis and pars tuberalis; although they appeared mostly as isolated cells, small groups of gonadotrophs were also identified in the pars distalis. In contrast, lactotrophs were observed only as clusters of cells exclusively in the pars distalis of sexually active and anoestrous mares in November and in most of the sexually active mares in July. A specific gonadotroph-lactotroph association was identified only between large isolated gonadotrophs and lactotroph clusters. Double immunofluorescent staining for FSHbeta and prolactin revealed a similar gonadotroph-lactotroph association to the one detected for LH gonadotrophs. No statistical difference in the gonadotroph:lactotroph ratio was observed as a result of changes in reproductive status or season. However, a tendency for a simultaneous decrease in the number of gonadotrophs and an increase in the number of lactotrophs was detected in anoestrous animals. Prolactin receptor immunoreactivity was found in the pars distalis, but not in the pars tuberalis, of sexually active (July and November) and anoestrous animals for both long and short forms of the receptor. No prolactin receptor co-localization for either form of the receptor was observed in LH or FSH gonadotrophs in either of the reproductive states examined during both summer and winter seasons. Furthermore, no significant difference was apparent in the proportion of cells expressing prolactin receptors between mares of different reproductive state or season. The specific anatomical association between gonadotroph and lactotroph cells and the expression of prolactin receptors in the equine pituitary gland indicate a potential role of prolactin in the regulation of gonadotrophin secretion. However, the absence of evidence for co-localization of prolactin receptors in LH or FSH cells does not support the hypothesis of a direct effect of prolactin on the gonadotroph as reported in a short day breeder. The results raise the possibility that, in horses, an intermediate regulatory cell may mediate the action of prolactin on gonadotroph function.     

Sequence analysis and expressional regulation of messenger RNAs encoding beta subunits of follicle-stimulating hormone and luteinizing hormone in the red-bellied newt, Cynops pyrrhogaster

Two distinct cDNAs encoding beta subunits of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were cloned from the cDNA library constructed for the pituitary of the red-bellied newt, Cynops pyrrhogaster, and sequenced. The newt FSHbeta and LHbeta cDNAs encode polypeptides of 129 and 131 amino acids, including signal peptides of 20 and 19 amino acids, respectively. The number and position of cysteine and N-glycosylation in each of the beta subunits of FSH and LH, which are considered essential for assembly of the alpha subunit, are well conserved between the newt and other tetrapods. The high homology (41.6%) between the beta subunits of newt FSH and LH imply less specificity of FSH and LH in gonadal function. One cDNA encoding the common polypeptide chain alpha subunit of FSH and LH was also isolated from the newt pituitary gland. The mRNAs of FSHbeta, LHbeta, and the alpha subunit were expressed only in the pituitary gland among various newt tissues. Double-staining with in situ hybridization and immunohistochemistry revealed coexpression of FSHbeta and LHbeta in the same newt pituitary cells. Ovariectomy induced a significant increase in FSHbeta mRNA levels, but there was no significant change in LHbeta or alpha subunit mRNA levels compared with those in control animals. Taken together, these data suggest that two kinds of gonadotropins, namely FSH and LH, are expressed in the same gonadotropin-producing cells in the pars distalis of the newt as well as in other tetrapods and that the expression of FSHbeta is negatively regulated by the ovaries.     

Dynamic changes in the gonadotrope cell subpopulations during an estradiol-induced surge in the ewe

Whether estradiol targets a subpopulation of gonadotrope cells was investigated in this study. Ovariectomized ewes (OVX) or OVX ewes immunized against GnRH and treated with hourly pulses of GnRH analogue (OVX-IMG) were killed at 6, 12, 16, and 24 h after administration of 50 microg of 17beta-estradiol (E(2)). Control ewes received no E(2) treatment. In OVX or OVX-IMG ewes killed 6 h after E(2) injection, a decrease in gonadotropin plasma levels was observed compared with non-E(2)-treated ewes. In contrast, a surge in gonadotropin plasma concentrations occurred in ewes killed 16 h after injection. The percentage of total immunoreactive gonadotrope cells among the pituitary cells was lower in E(2)-treated ewes compared with nontreated animals. The proportion of monohormonal LH cells was constant throughout the experiment, except at the surge peak, where it was enhanced. In the OVX ewes, the proportion of bihormonal LH/FSH cells was lower in the E(2)-treated ewes compared to the nontreated ewes (P: < 0.001), with a more pronounced decrease 16 h after E(2) injection. A slight increase occurred 12 h after E(2) injection compared with 6 h after injection (P: < 0.05). A similar pattern was observed in the OVX-IMG ewes, except at 12 h after E(2) injection, when no increase occurred. In both OVX and OVX-IMG ewes, injection of E(2) decreased FSHbeta mRNA expression but did not alter the relative levels of LHbeta mRNA. These data suggest that the negative feedback of E(2) on LH and FSH secretion mainly targets the bihormonal cells and occurs, at least in part, directly at the pituitary level. During the gonadotropin surge, the sustained FSH release from the bihormonal cells would induce a switch from bihormonal cells to monohormonal LH cells by depleting these cells of FSH.     

A comparative in vitro study on LHRH responsiveness of LH cells of the pars tuberalis and pars distalis

Summary The aim of the present study was to test whether the luteinizing-hormone (LH) cells in the pars tuberalis (PT) of the rat and mouse respond to LH-releasing hormone (LHRH) as do those of the pars distalis. A part of the basal hypothalamus containing the pituitary stalk, median eminence and the pars tuberalis (H-PT), was dissected out and incubated in vitro.The LH-secreting capacity of the PT was investigated after removal of the pituitary body (i.e., partes distalis, intermedia and nervosa). First, some rat and mouse H-PT tissues were treated with synthetic LHRH (100ng/ml), while others were incubated without LHRH. After 24 h of incubation, variable amounts of LH release were detected in the medium. This LH discharge, however, was not LHRH-dependent but proportional to the number of PT LH cells that were immunohistochemically detected in each incubated tissue. Since there was marked individual variation in the number of LH cells in the PT, the LH levels in the incubation medium were next compared before and after LHRH treatment using the same H-PT of the rat. An effect of LHRH could not clearly be shown in this experiment.Finally, the cytological response of the PT to LHRH was investigated by incubating both the H-PT and pituitary body connected to the intact pituitary stalk. Immunohistochemical examination of LHRH-treated tissues after 24 h revealed that, in females of both rats and mice, hormone depletion occurred in LH cells of the pars distalis but not in those of the PT. These results indicate that although LH cells in the PT can release LH in vitro, their mode of hormone synthesis and/or discharge differs from that of LH cells in the pars distalis. Since there was a marked individual variation and small LH-secreting capacity by the PT tissue, it seems unlikely, at least in rats and mice, that LH of PT origin plays an important role in the normal physiological state.     

Seasonal variations of gonadotropins in the pars distalis male viscacha pituitary. Effect of chronic melatonin treatment

The gonadotropes, LH and FSH cells, were immunohistochemically identified in the pituitary pars distalis of the adult male viscacha (Lagostomus maximus maximus) using specific antibodies against hLHbeta and hFSHbeta with the streptavidin-biotin-peroxidase complex. The distribution, size and percentage immunopositive area of these cells were analyzed by image analysis in viscachas captured during the annual reproductive cycle and after the chronic administration of melatonin. The LHbeta and FSHbeta cells showed seasonal changes in the distribution, size and percentage immunopositive area. The LHbeta cells were found widely distributed throughout the pars distalis during the reproductive period, and they were found in the ventro-medial region in the pars distalis during the gonadal regression and gonadal recovery periods. The LHbeta cells reached the largest size and immunopositive area during the reproductive period and the smallest size and immunopositive area during the gonadal regression period. The FSHbeta cells were found in the ventro-medial region during reproductive and gonadal regression periods. The FSHbeta cells were found widely distributed throughout the pars distalis during the gonadal recovery period when they showed the maximum percentage immunopositive area. A decrease in the size of LHbeta and FSHbeta cells was observed after the chronic administration of melatonin. Moreover, it produces a decrease in the immunopositive area occupied by the LHbeta cells but not in the immunopositive area occupied by the FSHbeta cells. Our results show great activity of LHbeta and FSHbeta cells in different moments of the annual reproductive cycle demonstrating that these cells do not secrete in parallel. Moreover, melatonin acts differentially on the activity of the gonadotrope cells.     

Subcellular localization of gonadotrophic hormones LH and FSH in frog adenohypophysis using double-staining immunocytochemistry

We applied double post-embedding immunocytochemical methods using specific antibodies against bullfrog (Rana catesbeiana) luteinizing hormone (LH) and follicle-stimulating hormone (FSH) with immunogold staining (5- and 20-nm particles) to determine the subcellular localization of both gonadotropins and to observe their immunostaining patterns in anterior pituitary of the frog Rana pipiens. Results showed that individual gonadotrophs may store either one or both gonadotropins in a given secretory granule and in large globules (lysosomes?). Most gonadotrophs (50-88%) contain both hormones; 12-50% contain only FSH, and only a few (0-7%) contain LH alone. Individual secretory granules, even in cells that contain both hormones, may contain only one or both gonadotropin molecules. Evaluation of the percentage of monohormonal and multihormonal secretory granules revealed that multihormonal secretory granules were the most numerous and that LH monohormonal secretory granules were the least numerous. These results indicate that cellular storage of gonadotropin in amphibian pituitary is similar to that described for mammals, where a single cell type containing both gonadotropins predominates. Variability in hormone content both of cells and of granules in all individuals is consistent with the hypothesis that frog pituitary possesses a single multipotential gonadotroph.     

Cellular associations of pituitary gonadotrophs in a rodent (Lagostomus maximus maximus) with photoperiod-dependent reproduction

The morphological characteristics and percentage of the cellular associations between gonadotrophs (LH- and FSH-secreting cells) and other cellular types were studied in pituitary pars distalis of adult male viscachas (Lagostomus maximus maximus) by double immunohistochemistry using specific antibodies to LH, FSH, PRL, GH, ACTH, TSH and S-100 protein (by folliculostellate cells; FSC), during long and short photoperiods. Bihormonal gonadotrophs were observed in ventro-medial and dorsal regions, interspersed between monohormonal gonadotrophs, and their number increased in short photoperiod. LH- and FSH-gonadotrophs were found around lactotrophs, enclosed by somatotrophs in the dorsal region, and associated with irregular corticotrophs. Gonadotrophs and thyrotrophs were associated along blood vessels and follicular structures. The cytoplasmic prolongations of FSC were in contact with both gonadotrophs. The percentage of LH–FSH, LH–ACTH, LH–FSC, FSH–LH, FSH–PRL, FSH–GH, FSH–ACTH, FSH–TSH and FSH–FSC associations decreased, whereas LH–PRL increased in short as compared to long photoperiod. The most abundant associations were LH–GH and LH–TSH during long photoperiod, but LH–GH and LH–PRL during short photoperiod. FSH–GH and FSH–PRL were the most numerous associations, and LH–FSC and FSH–FSC were the less abundant ones in both photoperiods. These results provide the morphological evidence for specific cellular associations between gonadotrophs and other cellular types of viscacha pituitary.     

Maintenance of gonadotrophs in pituitary autografts under the kidney capsules of female rats given sex hormones or LRH

Female Sprague-Dawley rats were hypophysectomized and the anterior pituitary gland was immediately placed under the kidney capsule. For 1 week after surgery, groups of pituitary autograft-bearing animals were treated with twice-daily injections of estradiol 17 beta (E), progesterone (P), estradiol 17 beta and progesterone (EP), or luteinizing hormone-releasing hormone (LRH). Within 2--4 hours following the last injection, the pituitary grafts were removed and placed into organ culture. They were maintained in culture with or without added LRH (10(-7) M) for 1 hour at 37 degrees C. The culture media were then frozen for later radioimmunoassay of FSH and LH. The tissues were kept in culture for an additional 24 hours, at which time they were fixed and prepared for immunocytochemistry or electron microscopy. Results showed that treatment of the animals with E, EP, or LRH enhanced the release of FSH and LH into the culture media, and that the release of these hormones was increased further by acute incubation with LRH. The ultrastructure of the gonadotrophs was well maintained by treating the animals with E or the combination of E and P or with LRH. Graft tissue from animals treated with LRH, which was incubated subsequently for 24 hours with LRH, showed the best maintenance of gonadotroph morphology. This experimental procedure should be useful for obtaining gonadotrophs for use in establishing gonadotroph cell lines.     

Induction of reproductive function in anestrous mares using a dopamine antagonist

We investigated the role of dopamine in the regulation of seasonal reproductive activity in mares. Nine seasonal anestrous mares, maintained under a natural photoperiod, were treated daily with a dopamine D2 antagonist, [-]-sulpiride (200 mg/mare, im), beginning February 5 (day of year = 36) until the first ovulation of the year or for a maximum of 58. Nine untreated anestrous mares were maintained under the same conditions. The ovaries were examined by ultrasonography twice a week, and blood was collected three times a week for progesterone, LH, FSH and prolactin determinations. Mean day of first ovulation was significantly advanced for [-]-sulpiride-treated mares than control mares (mean day of year +/- SEM = 77.3 +/- 7.9 and 110.0 +/- 6.8, respectively; P < 0.01). Eight mares ovulated during [-]-sulpiride treatment while one mare failed to ovulate. Ovulation occurred 91 d after the start of treatment or on Day 127. All mares continued to have normal estrous cycles after the first ovulation. First cycle length and luteal progesterone concentrations did not differ between [-]-sulpiride-treated and control mares. Plasma prolactin concentrations were significantly increased at 2 and 9 h after [-]-sulpiride administration (P < 0.05), and had returned to basal levels by 24 h. At the time of the LH surge associated with the first ovulation, mean LH and FSH secretion was significantly higher in [-]-sulpiride-treated mares than in control mares (P < 0.05). These results suggest that dopamine plays a role in the control of reproductive seasonality in mares and exerts a tonic inhibition on reproductive activity during the anovulatory season.     

Follicle-stimulating hormone pulse amplitude decreases with the onset of the breeding season in the mare

The relationship between daily mean FSH concentrations in serum and the pattern of FSH detected by frequent sampling for 12-h periods (samples every 15 min) was examined in five mares during the transition into the breeding season. The five mature anestrous mares were exposed to a natural increase in daylength. Blood samples were collected daily from February 1 until the first ovulation of the breeding season (April 14 +/- 3.7 days, Mean +/- SEM). Periods of frequent blood collection were performed every two weeks. Blood samples were obtained daily by jugular venipuncture or jugular cannula (frequent samples). Mean daily concentrations of FSH in serum determined by RIA decreased during seasonal transition. Patterns of FSH in serum detected by frequent sampling were pulsatile. FSH pulse amplitude decreased during seasonal transition, and the decrease in amplitude was associated with the decrease in mean serum FSH concentrations. This decrease in FSH pulse amplitude may reflect an involvement of a follicular product from developing follicles or a change in hypothalamic stimulation of pituitary FSH release.     

Localization of an endocannabinoid system in the hypophysial pars tuberalis and pars distalis of man

The hypophysial pars tuberalis (PT) acts as an important interface between neuroendocrine brain centers (hypothalamus, pineal organ) and the pars distalis (PD) of the hypophysis. Recently, we have identified an endocannabinoid system in the PT of hamsters and provided evidence that 2-arachidonoylglycerol is a messenger molecule that appears to play an essential role in seasonal reproduction and prolactin release by acting on the cannabinoid receptors in the PD. We now demonstrate the enzymes involved in endocannabinoid synthesis and degradation, namely sn-1-selective diacylglycerol lipase α, N-acylphosphatidylethanolamine-specific phospholipase D, and monoacylglycerol lipase, in the PT of man by means of immunohistochemistry. High-performance liquid chromatography coupled with tandem mass spectrometry revealed 2-arachidonoylglycerol and other endocannabinoids in the human PT. Furthermore, we detected the expression of the cannabinoid receptor 1 (CB1), a primary receptor for endocannabinoids, in the PD. Double-immunofluorescence staining for CB1 and various hypophysial hormones or S-100, a marker for folliculostellate (FS) cells, revealed that CB1 immunoreactivity was mainly localized to corticotrophs and FS-cells. A limited number of lactotrophs and somatotrophs also showed CB1 immunoreactivity, which was however absent from gonadotrophs and thyrotrophs. Our data thus indicate that the human PT comprises an endocannabinoid system, and that corticotrophs and FS-cells are the main target cells for endocannabinoids. The functional significance of this newly discovered pathway remains to be elucidated in man; it might be related to the control of stress responses and/or reflect a remnant seasonal control of hypophysial hormonal secretion.     

Phosphatidic acid and the calcium-dependent actions of gonadotropin-releasing hormone in pituitary gonadotrophs

The stimulation of luteinizing hormone (LH) release and cyclic GMP (cGMP) production in rat anterior pituitary cells by gonadotropin-releasing hormone (GnRH) are receptor mediated and calcium dependent, and have been shown to be accompanied by increased phospholipid turnover and arachidonic acid release. The incorporation of 32Pi into the total phospholipid fraction of pituitary gonadotrophs was significantly elevated by 10(-8) M GnRH, with specific increases in the labeling of phosphatidylinositol and phosphatidic acid (PA). Since PA acts as a calcium ionophore in several cell types, its effects upon calcium-mediated gonadotroph responses were compared with those elicited by GnRH. In rat pituitary gonadotrophs prepared by centrifugal elutriation, PA stimulated LH release and cGMP production by 9-fold and 5-fold, respectively. The stimulation of LH release by 30 microM PA was biphasic in its dependence on extracellular calcium concentration, rising from zero in the absence of calcium to a maximum of 10-fold at 0.5 mM Ca2+ and declining at higher calcium concentrations. In dose-response experiments, PA was 3-fold more potent at 0.5 mM Ca2+ than at 1.2 mM Ca2+. The cGMP response to PA in cultured gonadotrophs was also calcium dependent, and was progressively enhanced by increasing Ca2+ concentrations up to 1.5 mM. The ability of PA to stimulate both LH release and cGMP formation in a calcium-dependent manner suggests that endogenous PA formed in response to GnRH receptor activation could function as a Ca2+ ionophore in pituitary gonadotrophs, and may participate in the stimulation of gonadotroph responses by GnRH and its agonist analogs.     

Changes in the hypothalamic-hypophyseal axis of mares associated with seasonal reproductive recrudescence

Four groups of mares, representing anestrus (AN; n = 8), early transition (ET; n = 7), late transition (LT; n = 8) and estrus (EST; n = 12) were used to examine changes in the hypothalamus and anterior pituitary during the period of transition from winter anestrus into the breeding season. Mares were of mixed breeding, between the ages of 3 and 20 years, and had shown normal patterns of estrous behavior and ovulation during the breeding season previous to this experiment. Hypothalamic content of gonadotropin-releasing hormone (GnRH) and anterior pituitary content of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were determined by radioimmunoassay. The number of receptors for GnRH in anterior pituitary tissue was also determined. There was no effect of stage of transition into the breeding season on receptors for GnRH or content of FSH (p greater than 0.05). Likewise, content of GnRH in the hypothalamus did not differ between the four groups (p greater than 0.05). However, pituitary content of LH increased progressively from anestrus to the breeding season (p less than 0.05). Means for the AN, ET, LT and EST groups were 1.1 +/- 0.2, 2.2 +/- 0.3, 6.3 +/- 1.4 and 15.2 +/- 1.8 micrograms LH/mg pituitary, respectively. In addition, serum concentrations of LH associated with the first ovulation of the year for 5 of the EST mares were significantly lower (p less than 0.01) than those associated with the second ovulation of the year.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immunocytochemistry of the pituitary glycoprotein hormones.

The storage sites of the pituitary glycoprotein hormones were identified with the use of electron microscopic immunocytochemical techniques and antisera to the beta (beta) chains of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and thyroid-stimulating hormone (TSH). The TSH cells in normal rats is ovoid or angular and contains small granules 60-160 nm in diameter. In TSH cells hypertrophied 45 days after thyroidectomy, staining is in globular patches in granules or diffusely distributed in the expanded profiles of dilated rough endoplasmic reticulum. The gonadotrophs (FSH and LH cells) exhibited three different morphologies. Type I cells are ovoid with a population of large granules and a population of small granules. Staining for FSHbeta or LHbeta was intense and specific only in the large granules (diameter of 400 nm or greater). Type II cells are angular or stellate and contain numerous secretory granules averaging 200-220 nm in diameter. They predominate during stages in the estrous cycle when FSH or LH secretion is high. Type III cells look like adrenocorticotropin (ACTH) cells in that they are stellate with peripherally arranged granules. They generally stain only with anti-FSHbeta and their staining can not be abolished by the addition of 100 ng ACTH. In preliminary quantitative studies of cycling females, we found that on serial sections FSH cells and LH cells show similar shifts to a more angular population of cells during stages of active secretion. However, the shifts are not in phase with one another. Furthermore, there are at least 1.5 times more FSH cells than LH cells at all stages of the cycle. Our collection of serial cells shows that some cells (usually type I or II) stain for both gonadotropic hormones, whereas others (usually type II or III) contain only one.     

Changes in the hypothalamic-hypophyseal axis of mares in relation to the winter solstice.

In mares, the amount of gonadotrophin-releasing hormone (GnRH) is low in the hypothalamus during seasonal anoestrus, but by early spring, concentrations of GnRH are high. The timing of this response was characterized more precisely by determining concentrations of GnRH in hypothalamic tissue collected immediately before and at various times after the winter solstice (22 December 1986). Ovaries, pituitary gland, hypothalamus and a blood sample were collected from six groups of mares (6-12 mares per group) at death, 1 week before day of the winter solstice and 1, 2, 3 and 12 weeks afterwards. No significant changes in weight of the anterior pituitary gland or concentrations of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were observed in the anterior pituitary gland (P > 0.1). Mean diameter of the largest follicle, number of follicles > or = 20 mm in diameter and concentrations of LH and FSH in serum remained unchanged for weeks -1 to +3 (P < 0.05), then increased significantly by week 12 (P < 0.001). Content and concentration of GnRH in the median eminence was low at -1 week, increased gradually (P < 0.05) to a maximum by +1 week, then decreased gradually (P < 0.05) to low values at 12 weeks. Means (+/- SEM) for -1, +1 and +12 weeks were 33.5 +/- 5.5, 117.7 +/- 18.6 and 29.8 +/- 3.7 ng GnRH, respectively. Mean content of GnRH in the preoptic area of the hypothalamus showed a reciprocal pattern.(ABSTRACT TRUNCATED AT 250 WORDS)     

Gonadotropin-releasing hormone treatment induces follicular growth and ovulation in seasonally anestrous mares

A study was conducted to evaluate the effectiveness of gonadotropin-releasing hormone (GnRH) pulse infusion to stimulate follicular development and induce ovulation in seasonally anestrous standardbred mares. Seventeen mares were selected for use in this experiment, on the basis of a previous normal reproductive history, and were housed under a photoperiod of 8L:16D beginning one week prior to the start of the experiment (second week in January). Mares were infused with 20 micrograms (n = 7) or 2 micrograms (n = 6) GnRH/h, or were subjected to photoperiod treatment only (controls, n = 4). Serum concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and progesterone did not vary, and neither significant follicular development nor ovulation was observed in any control mare throughout the experimental period (greater than 60 days). By contrast, both groups of GnRH-treated mares showed elevated serum concentrations of LH and FSH within one day after the start of infusion. Mares infused with 20 micrograms GnRH/h had at least one follicle greater than or equal to 25 mm in 7.4 +/- 1.3 (mean +/- SEM) days following the start of infusion, and ovulated in 12.0 +/- 0.7 days. In the 2-microgram-GnRH/h treatment group, a 25-mm follicle was detected in 5.7 +/- 0.7 days, and ovulation occurred after 10.0 +/- 0.3 days of infusion. Ovulation in every instance was followed by a functional luteal phase, as indicated by the profiles of progesterone secretion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immunoreactivity of gonadotrophs (FSH and LH Cells) and gonadotropin subunit gene expression in the male chub mackerel Scomber japonicus pituitary during the reproductive cycle

The gonadotropins (GtHs), follicle-stimulating hormone (FSH) and luteinizing hormone (LH), are heterodimers composed of a common α subunit (GPα) and a unique β subunit (FSHβ or LHβ); they are synthesized in and secreted from gonadotrophs (FSH and LH cells) in the pituitary. Little is known about the roles of FSH and LH during spermatogenesis in perciform fishes. In this study, we examined immunoreactive changes in FSH and LH cells, and changes in the gene expression of the three gonadotropin subunits in the pituitary of male chub mackerel Scomber japonicus during testicular development. FSHβ-immunoreactive (ir) and LHβ-ir cell area were measured immuno-histochemically based on the FSH and LH cell-occupying area in the proximal pars distalis. The FSHβ-ir cell area increased significantly during spermiation, while FSHβ mRNA levels, already high at the beginning of spermatogenesis, increased further, peaking during spermiation. In contrast, LHβ-ir cell area and LHβ mRNA levels, which were low at the beginning of spermatogenesis, increased significantly during late spermatogenesis, peaking during spermiation. For both FSH and LH, GtHβ-ir cell area and GtHβ mRNA levels decreased until gonadal resting. GPα mRNA levels showed similar changes to LHβ mRNA levels. These results suggest that in the chub mackerel, FSH may play an important role in the early and late phases of spermatogenesis, and that LH may play a role during late spermatogenesis and spermiation. Moreover, our results demonstrate that changes in GtHβ-ir cell area were accompanied by similar changes in the expression of the FSHβ and LHβ genes, both of which increased during testicular development.