Reproduction in the Mexican leaf frog, Pachymedusa dacnicolor. VI. Presence and distribution of multiple GnRH forms in the brain.

The presence and distribution of gonadotropin-releasing hormone (GnRH) has been investigated in the Mexican leaf frog, Pachymedusa dacnicolor, brain during development and in the adult. The ontogenetic pattern of GnRH neurons illustrates their extracranial as well as intracranial sites. Immunohistochemical analysis indicates that GnRH-immunoreactive neurons appear during the metamorphic climax. They are located in the mesencephalon and subsequently other GnRH neurons appear in the peripheral terminal nerve and anterior preoptic area of the brain. Use of specific antisera and homologous combined with heterologous preabsorption tests indicate that mammalian and chicken GnRH-II-like peptide-containing neurons are differentially located within the brain, the former in the anterior preoptic area and peripheral terminal nerve and the latter in the mesencephalon. HPLC and RIA data suggest the presence of three forms of immunoreactive GnRH in the P. dacnicolor brain. A mammalian GnRH-like molecule and a chicken GnRH-II-like molecule are present. A third form, suspected to be [hydroxyproline9]mGnRH elutes before the mammalian GnRH.

Influence of gonadectomy and steroid hormone replacement therapy on the gonadotropin-releasing hormone neuronal system in the anterior preoptic area of the frog (Rana esculenta) brain.

Effects of gonadectomy and sex steroid replacement therapy were studied on the anterior preoptic area-located GnRH-immunoreactive neurons in males and females of the frog, Rana esculenta. Removal of gonads caused a drastic reduction in the immunostaining as well as in the number of GnRH neurons in the anterior preoptic area of the brain in both sexes. Replacement therapy with estradiol-17 beta, testosterone, and 5 alpha-dihydrotestosterone, given alone or in combination, enhanced the somal accumulation of immunoreactive material in GnRH neurons and also their number. These effects, however, varied to some extent from steroid to steroid, a combination of testosterone and 5 alpha-dihydrotestosterone being most effective in gonadectomized males, whereas that of estradiol-17 beta and testosterone was most effective in gonadectomized females. The present data are the first to demonstrate that androgen and estrogen can influence the immunoreactive GnRH neurons in the anterior preoptic area of the amphibian brain.

Immunohistochemical localization of GnRH in the crested newt (Triturus carnifex) brain and terminal nerve: a developmental study.

Localization of GnRH-immunoreactive neuronal system was studied by immunohistochemistry in the nasal-brain area of the crested newt, Triturus carnifex. Besides adults, developmental stages were those from hatchlings up to complete metamorphosis. Neurons containing immunoreactive GnRH were first detected in the nasal area of larvae with yet undifferentiated gonads. Subsequently, in prometamorphic stages, GnRH-immunoreactive cell bodies and fibers were detected in the proximal part of the terminal nerve as well as along the ventromedial surface of the olfactory bulbs. In older larvae with sexually differentiated gonads and up to the metamorphic climax GnRH-neurons were detected, as a rostral to caudal continuum, along the ventromedial surface of the olfactory bulbs and midtelencephalon. This is exactly the route followed by the terminal nerve. In the adult brain, besides the presence of occasional GnRH-neurons and fibers in the terminal nerve proximal to olfactory bulbs, olfactory bulbs and the mid-basal telencephalon, another aggregate of immunoreactive neurons was present in the anterior preoptic area, and a greater number of fibers in the habenular area as well as in the infundibular floor, median eminence and pars nervosa. These data suggest the nasal area to forebrain migration (along the course of the terminal nerve) of GnRH-neurons during development in the crested newt.

Immunohistochemical demonstration of the presence and localization of diverse molecular forms of gonadotropin-releasing hormone in the lizard (Podarcis s. sicula) brain.

The immunohistochemical presence and the distribution pattern of four different molecular forms of gonadotropin-releasing hormone (GnRH) were investigated in the brain of both sexes of the lizard, Podarcis s. sicula. Animals used in this study were collected in November and April, representing two different periods of the reproductive cycle. The antisera used were those raised against synthetic mammalian GnRH, chicken GnRH-I and II, and salmon GnRH. Strong immunoreaction was obtained for salmon, chicken-I, and chicken-II GnRHs, whereas a very weak reaction was seen for the mammalian form of GnRH. The distribution of immunoreactive-GnRH perikarya and fibers did not vary with the sex, the reproductive condition of the animals, or the antiserum used. Also, the intensity of immunoreaction with any one antiserum was quite similar in both periods of the year and in all brains examined. The immunoreactive perikarya was seen as two distinct groups, one in the mesencephalon and the other in the infundibulum. Immunoreactive fiber endings were seen in the telencephalon, the optic tectum, the anterior preoptic area, the median eminence, the central grey matter, the rhombencephalon, and the cerebellum. No immunoreactive perikarya were seen in the telencephalon or the anterior preoptic area.

Reproduction in the Mexican leaf frog, Pachymedusa dacnicolor. V. Immunohistochemical localization of gonadotropin-releasing hormone in the brain.

The presence and distribution of immunoreactive gonadotropin-releasing hormone (GnRH) in brains of adult male and female Pachymedusa dacnicolor has been studied immunohistochemically using antisera against mammalian, chicken-II, and salmon GnRHs. The distribution map of the immunoreactive-GnRH elements in the brain of P. dacnicolor is extremely simple, being limited to the anterior preoptic area-infundibulum-median eminence circuit. No sex- or reproductive status-related difference in either the distribution pattern or intensity of immunoreaction was revealed in this study. This is also the first immunohistochemical evidence of the presence of different structural forms of GnRH in the brain of an amphibian.

Immunoreactive luteinizing hormone-releasing hormone in the frog (Rana esculenta) brain: distribution pattern in the adult, seasonal changes, castration effects, and developmental aspects.

The present work describes the neuroanatomical distribution of the immunoreactive luteinizing hormone-releasing hormone (ir-LHRH) system in the brain of adult male and female, castrated male and developing Rana esculenta. No obvious sex differences in the distribution pattern of ir-LHRH were observed. Immunoreactive neuronal cell bodies are not contained within a single anatomically defined area of the brain. They are present as distinct groups in the olfactory bulbs, medial septal area, anterior preoptic area (APOA), retrochiasmatic area of the infundibulum, and interpeduncular nucleus-tegmentum area. Of the entire brain, the medial septal-APOA region exhibits the highest frequency of ir-LHRH cell bodies in both sexes. ir-LHRH fiber projections are present in the olfactory bulbs, medial septal area, APOA, floor of the diencephalon, subhabenular-periventricular area in the epithalamus, lateral suprachiasmatic area, ventrolateral infundibulum, median eminence, pars nervosa, optic tectum, interpeduncular nucleus-tegmentum area, and rhombencephalon grey. Castration seems to bear no effect on the pattern of ir-LHRH system in the frog brain. The influence of castration consisted in decreased intensity of the immunostaining and frequency of occurrence of the septal-APOA neuronal cell bodies. In median eminence, castration also induced a sensible decrease in the immunoreactivity, whereas in the pars nervosa of 50-day castrates ir-LHRH fibers totally disappeared. During ontogenesis, ir-LHRH elements first become evident in stage 31 tadpoles (beginning of metamorphic climax); LHRH immunoreaction is restricted to the cell bodies and fibers in the APOA and some fibers in the ventral hypothalamus and a few in median eminence. This condition remains unaltered until stage 33 when the tail is almost totally resorbed. The possible implications of the ir-LHRH-containing brain areas in the different aspects of reproduction in the frog are briefly discussed.

Reproduction in the Mexican leaf frog, Pachymedusa dacnicolor. II. The male.

The Mexican leaf frog, Pachymedusa dacnicolor, an inhabitant of the semiarid, subtropical Mexican lowlands, displays a well-defined seasonal testicular cycle. Testis weight seems to be a reliable index of the reproductive status of the animal and plasma levels of androgens (testosterone, T; 5 alpha-dihydrotestosterone, DHT; androstenedione, A) correlate not only with testicular growth, but with callosity development, reproductive behavior, and breeding. During the fall and winter, testis weight reaches its minimum as do plasma concentrations of T, DHT, A, and estradiol-17 beta (E). Plasma levels of progesterone (P) are maintained at a very low level throughout the year. During the fall and winter, spermatogenesis is almost entirely absent and the callosities are white and smooth. No signs of reproductive behavior (calling and amplexus) are evident. Late spring marks the initiation of spermatogenesis, testis weight increase, darkening of callosities, and increase in plasma levels of androgens. The magnitude of callosity development and the onset of calling and amplectant behavior are correlated with a great rise in plasma androgen levels. Although plasma T concentrations were higher than plasma levels of DHT and A, this androgen failed to stimulate the development of callosities and calling behavior in successfully castrated males.

Seasonal testosterone profile and testicular responsiveness to pituitary factors and gonadotrophin releasing hormone during two different phases of the sexual cycle of the frog (Rana esculenta).

Plasma and testicular testosterone concentrations in the frog, Rana esculenta, were studied by radioimmunoassay and showed similar seasonal fluctuations. The increase in testicular androgen during November preceded that occurring in the plasma by 2 months. Pituitary products and gonadotrophin releasing hormone, and the responsiveness of the testis to these substances play an important role in determining the hormone profile.

Seasonal plasma sex steroid levels in the female Rana esculenta.

Seasonal plasma progesterone, androstenedione, estrone, and 17 beta-estradiol concentrations in the female Rana esculenta were determined by radioimmunoassay during the 1979 and 1981 seasons. Plasma levels of these steroids were highest just before the first ovulatory wave in spring and lowest after the breeding season. In the 1979 season (during the 1981 season hormones were not assayed in January, November, and December) progesterone, androstenedione, and estradiol levels showed another peak in November-December. During the breeding months, i.e., late March to late June, progesterone, androstenedione, and estradiol levels showed intermittent ups and downs corresponding roughly to the ovulatory waves. In addition, during the breeding season progesterone and androstenedione levels had a higher average in frogs with "ripe" ovaries than in those with "spent" ovaries. Relationships between seasonal steroid levels and ovarian activity are briefly discussed.

Effects of anesthesia, surgical manipulation and dehydration on the nucleic acid and protein content of the pituitary and hypothalamus of the frog.

Various types of stress, chemical or surgical, have a negative influence on the protein and RNA content of the hypothalamo-hypophyseal system of Rana esculenta. A degree of recovery occurs in these tissues 24 h after MS222 anaesthesia and laparotomy. Decapitation is apparently the most suitable method of sacrificing the animals.

The function of fat bodies in relation to the hypothalamo-hypophyseal-gonadal axis in the frog, Rana esculenta.

In this study the authors have tried to furnish experimental support for the importance of fat bodies in the normal functioning of the hypothalamo-hypophyseal-gonadal system of the male frog, Rana esculenta. These experiments have shown a hypothalamo-hypophyseal control of the mobilization of fat body contents, directly involved in the control of testicular activity. Furthermore it is proposed that the fat body contents are released into the testis through direct vascular contacts between the two organs. We suggest that the A1 cells (lactotrophs) and/or B2 cells (FSH-gonadotrops) of the pars distalis gonadotropins are incapable of stimulating the testis in the absence of fat bodies. In the light of these results a scheme has been put forward showing the position of fat bodies in the hypothalamo-hypophyseal-gonadal axis of the frog.