Hypertrophy of gonadotropin releasing hormone-containing neurons after castration in the teleost, Haplochromis burtoni.

In the African cichlid fish, Haplochromis burtoni, males are either territorial or nonterritorial. Territorial males suppress reproductive function in the nonterritorial males, and have larger gonads and larger gonadotropin-releasing hormone- (GnRH) containing neurons in the preoptic area (POA). We describe an experiment designed to establish the causal relationship between large GnRH neurons and large testes in these males by determining the feedback effects of gonadal sex steroids on the GnRH neurons. Territorial males were either castrated or sham-operated, 4 weeks after which they were sacrificed. Circulating steroid levels were measured, and the GnRH-containing neurons were visualized by staining sagittal sections of the brains with an antibody to salmon GnRH. The soma areas of antibody-stained neurons were measured with a computer-aided imaging system. Completely castrated males had markedly reduced levels of circulating sex steroids [11-ketotestosterone (11KT) and testosterone (T)], as well as 17 beta-estradiol (E2). POA GnRH neurons in castrates showed a significant increase in mean soma size relative to the intact territorial males. Hence, in mature animals, gonadal steroids act as a brake on the growth of GnRH-containing neurons, and gonadal products are not responsible for the large GnRH neurons characteristic of territorial males.     

Gonadectomy,sex hormones and the growth of tetrathyridial populations of Mesocestoides corti (Cestoda: Cyclophyllidea) in mice

Gonadectomy in mice considerably depressed the volumes of intraperitoneal tetrathyridial populations, orchiectomy being more efficient than ovariectomy. Testosterone propionate accelerated the growth of tetrathyridial populations in gonadectomized mice of both sexes. Oestradiol benzoate was less efficient, but considerably increased the invasion of livers both in intact and in gonadectomized mice. The average size of tetrathyridia was inversely proportional to the size of the populations: tetrathyridia from an orchiectomized mouse were the largest, those from an orchiectomized and testosterone treated mouse, the smallest, and those from an orchiectomized mouse treated with oestradiol, intermediate in size.     

Effect of intrauterine administration of gonadotropin releasing hormone on serum LH concentrations in lactating dairy cows

The objectives were to compare: (1) preovulatory serum LH concentrations, and (2) synchronization of ovulation, after im or iu administration of the second GnRH treatment of Ovsynch in lactating dairy cows. Lactating cows (N = 23) were presynchronized with two injections of PGF_2α given 14 days apart (starting at 34 ± 3 days in milk), followed by Ovsynch (GnRH-7 d-PGF_2α-56 h-GnRH) 12 days later. At the time of the second GnRH of Ovsynch (Hour 0), cows were blocked by parity and randomly assigned to 1 of 3 groups: (1) control group (CON; N = 7) were given 2 mL sterile water im; (2) intramuscular group (IM; N = 8) received 100 μg of GnRH im; and (3) intrauterine group (IU; N = 8) had 100 μg GnRH infused in the uterus (2 mL). Blood samples for serum LH concentrations were collected at Hours 0, 0.5, 1, 1.5, 2, 3, and 4. Furthermore, ultrasonography was performed twice daily (12-h intervals) from Hours 0 to 60 to confirm ovulation. The LH concentrations were greater (P < 0.05) in the IM than IU and CON groups at Hours 0, 0.5, 1, 1.5, 2, 3, and 4. Although LH concentrations were numerically higher in the IU group, LH concentrations within the IU and CON groups did not change over time. More cows ovulated in the IM (8/8) and IU (7/8) groups within 60 h after the second GnRH administration compared with the CON (2/7) group. In summary, serum LH concentrations were lower in the IU versus IM group, but the proportion of cows that ovulated within 60 h was similar between these two groups. Therefore, iu administration of GnRH may be an alternative route of delivery to synchronize ovulation in beef and dairy cattle.     

Ipsilateral retinal projections into the tectum during regeneration of the optic nerve in the cichlid fish Haplochromis burtoni: A dil study in fixed tissue

Retinal projections were experimentally manipulated in a bony fish to reveal conditions under which considerably enlarged ipsilateral projections developed and persisted. Three experimental groups were studied: animals after unilateral enucleation, after unilateral nerve crush, and after enucleation and crush of the remaining optic nerve. At 29 days after unilateral enucleation alone, no enhanced ipsilateral projection had developed. After nerve crush, however, large numbers of retinal fibers regenerated into the ipsilateral tectum. Retrogradely filled, ipsilaterally projecting ganglion cells were distributed throughout the entire retina. After 15 days regenerating retinal fibers covered the entire ipsilateral tectum. At later stages the ipsilateral projection showed progressive reduction in coverage of the tectum. Combining enucleation with nerve crush led to an ipsilateral projection that covered the tectum at 28 days and later. In this experimental situation the development of an ipsilateral projection appears to be a two-step process: (1) Fibers are rerouted to the ipsilateral side at the diencephalon, and (2) ipsilateral fibers persist in the tectum only in the absence of a contralateral projection while they appear to be eliminated in the other cases. © 1992 John Wiley & Sons, Inc.     

Effect of clomiphene citrate on pituitary responsiveness to gonadotropin releasing hormone in rams and wethers

The objective of this study was to determine the effect of clomiphene citrate (clomid) on pituitary responsiveness to gonadotropin releasing hormone (GnRH) in rams and wethers. Doses of 200 mg clomid per ram and 1 mug GnRH per 50 kg body weight were used in studies on 12 rams and 4 wethers. The experimental design involved bleeding each animal at 15-minute intervals for 6.5 hours. At the end of the first hour, GnRH was injected IV. The second GnRH challenge was administered 0.5 hours after an injection of clomid or vehicle (4.5% sorbitol solution) which was given on the third hour. The relative response to clomid or vehicle was calculated as the mean increase in concentration of LH during the two-hour period after the second GnRH injection. Each treatment (clomid and vehicle) was given to all animals with a 14-day recovery period between treatment days. The relative response for the rams receiving vehicle (1.80 +/- 0.65) was greater (P < 0.05) than the response during clomid treatment (0.34 +/- 0.22). This suppression of LH response by clomid was observed in 10 of the 12 rams. In contrast to the rams, the concentrations of LH in wethers after the second GnRH injection were lower than those observed after the first GnRH injection. Similar to the rams, the relative response following clomid treatment of wethers (0.04 +/- 0.04) was less than the relative response (P > 0.05) following vehicle (0.40 +/- 0.16). The results suggest that clomid at this dosage inhibits GnRH-induced release of LH from the pituitary of rams but not of wethers.     

Plasma luteinizing hormone concentrations in cows given repeated treatments or three different doses of gonadotropin releasing hormone

We hypothesized that: (i) repeated GnRH treatments would increase the magnitude and duration of the LH surge and would increase progesterone (P4) concentrations after ovulation; and (ii) the release of pituitary LH would be greater in response to larger doses of GnRH. In Experiment 1, ovary-intact cows were given an intravaginal P4 (1.9 g) insert (CIDR) for 10 d and 500 μg cloprostenol (PGF) at CIDR removal to synchronize estrus. On Days 7 or 8 after estrus, cows received two PGF treatments (12 h apart) and 100 μg GnRH at 36 (Control), 36 and 38 (GnRH38), or 36 and 40 h (GnRH40) after the first PGF. Mean plasma LH concentration (ng/mL) was greater (P < 0.05) in GnRH38 (8.8 ± 1.1) than in Control (5.1 ± 1.3), with that in GnRH40 (5.8 ± 1.3) being intermediate. Although the duration (h) of the LH surge was longer in GnRH40 (8.0 ± 0.4) than in either GnRH38 (P < 0.05; 7.0 ± 0.3) or Control (P < 0.09; 7.1 ± 0.4), mean postovulatory P4 (ng/mL) was greater (P < 0.01) in Control (4.2 ± 0.7) than in GnRH38 (2.9 ± 0.6) or GnRH40 (3.0 ± 0.7) cows. In Experiment 2, ovariectomized cows were given a CIDR for 10 d and 2 mg of estradiol cypionate im at CIDR insertion. Thirty-six hours after CIDR removal, cows received, 50, 100, or 250 μg of GnRH. Cows given 250 μg GnRH released more LH (9.4 ± 1.4 ng/mL) than those given 50 or 100 μg (6.1 ± 1.3 and 5.4 ± 1.4 ng/mL, respectively), and had an LH surge of longer duration than those given 50 μg (6.8 ± 0.4 vs. 5.1 ± 0.3 h). In summary, ovary-intact cows in the GnRH38 group had greater mean and peak LH concentrations, but subsequent plasma P4 concentrations were lower than in Control cows. Ovariectomized cows given 250 μg GnRH had a greater pituitary release of LH.     

Dendrites determine the contribution of after depolarization potentials (ADPs) to generation of repetitive action potentials in hypothalamic gonadotropin releasing-hormone (GnRH) neurons

The impact of structure in modulating synaptic signals originating in dendrites is widely recognized. In this study, we focused on the impact of dendrite morphology on a local spike generating mechanism which has been implicated in hormone secretion, the after depolarization potential (ADP). Using multi-compartmental models of hypothalamic GnRH neurons, we systematically truncated dendrite length and determined the consequence on ADP amplitude and repetitive firing. Decreasing the length of the dendrite significantly increased the amplitude of the ADP and increased repetitive firing. These effects were observed in dendrites both with and without active conductances suggesting they largely reflect passive characteristics of the dendrite. In order to test the findings of the model, we performed whole-cell recordings in GnRH neurons and elicited ADPs using current injection. During recordings, neurons were filled with biocytin so that we could determine dendritic and total projection (dendrite plus axon) length. Neurons exhibited ADPs and increasing ADP amplitude was associated with decreasing dendrite length, in keeping with the predictions of the models. Thus, despite the relatively simple morphology of the GnRH neuron’s dendrite, it can still exert a substantial impact on the final neuronal output. This work was supported by HD-45436 to KJS and by NCRR P20 RR16481 to Nigel Cooper.     

The angular orientation of the black eye-bar in Haplochromis burtoni (Cichlidae,Pisces) and its relevance to aggressivity

Summary The effect of the black eye-bar in Haplochromis burtoni on the attack rate of conspecific males was found, using dummies, to depend on the angular orientation of this bar with respect to the body axis of the fish (Figs. 1, 4): The increment in attack rate increases as the black eye-bar parallels more closely the profile of the forehead and decreases as the bar approaches an orientation perpendicular to the forehead. This is valid independent of the actual body posture of the dummy bearing the black bar as could be shown by presenting dummies in a horizontal as well as in a head downward position. However, the head-down posture itself was found to cause an extra increment in attack rate, which may be considered to be additively or multiplicatively superimposed upon the increment determined by the orientation of the eye-bar with respect to the body axis (Fig. 4).Mit Unterstützung der Deutschen Forschungsgemeinschaft im Rahmen des SFB 50.We wish to thank Ann and John Thorson for their most stimulating discussion of the paper.     

Scanning electron microscopy of photoreceptor cells in the light- and dark-adapted retina of Haplochromis burtoni (Cichlidae,Teleostei)

Summary The photoreceptor layer in the retina of Haplochromis burtoni (Cichlidae, Teleostei) was studied by scanning electron microscopy. Three types of receptors were identified: rods, single-cones and double-cones. The three-dimensional arrangement of these photoreceptors is described in the light- and dark-adapted retina. The surface of the inner segment of the photoreceptor cells displays fine vertical fissures which give rise to slender processes. These so called calycal processes which are of different lengths in rods and cones, surround the beginning of the smooth-surfaced outer segment. The myoid, the contractile part of the receptor, which is located beneath the ellipsoid, was examined in the single-cones of the dark-adapted retina. It is a slender structure with surface infoldings. The myoid, studied by transmission electron microscopy, contains bundles of parallel myofilaments, which are thought to be contractile.This investigation was supported by grants of the Deutsche Forschungsgemeinschaft (Sonderforschungsbereich 51-E/10)     

Effects of testosterone, dihydrotestosterone and estradiol on gonadotropin release after gonadotropin releasing hormone administration in cyclic mares

Sixteen intact cyclic mares were treated on the fourth day of estrus and then every other day for a total of six injections with 1) testosterone propionate, 2) dihydrotestosterone (DHT) benzoate, 3) estradiol (E2) benzoate or 4) safflower oil. Mares were given gonadotropin releasing hormone (GnRH) on Day 3 of estrus (pretreatment) and again 24 h after the last steroid or oil injection. Treatment with testosterone propionate resulted in a greater (P less than 0.05) follicle-stimulating hormone (FSH) response to the second injection of GnRH compared with all other treatments. Treatment with DHT benzoate also resulted in greater (P less than 0.05) FSH response to GnRH compared with control and E2 benzoate-treated mares. Testosterone propionate and E2 benzoate administration suppressed (P less than 0.05) the normal diestrous rise in FSH concentrations exhibited by the control and DHT benzoate-treated mares. Steroid treatment did not affect the luteinizing hormone (LH) response to GnRH, although testosterone propionate treatment did suppress concentrations of LH in daily blood samples during Days 3 to 6 of treatment. It is concluded that testosterone's effect on FSH after GnRH treatment observed in this and previous experiments can be attributed to two different properties of the hormone or its metabolites acting simultaneously. That is, testosterone increased the secretion of FSH in response to GnRH as did DHT (an androgenic effect). At the same time, testosterone suppressed FSH concentrations in daily blood samples in a manner identical to that of E2 benzoate (an estrogenic effect).     

Effects of 72 hr calf removal and/or gonadotropin releasing hormone on luteinizing hormone release and ovarian activity in postpartum beef cows

A study was conducted to determine the pituitary and ovarian responses to 72 hr calf removal (CR) and/or gonadotropin releasing hormone (GnRH) in beef cows. Forty-eight Angus, Simmental, and Charolais crossbred cows in moderate body condition were allotted to an experiment of 2 x 2 factorial design involving CR and GnRH. At 30 to 32 days postpartum, calves were removed for 72 hr from the CR and CR plus GnRH groups. All cows were injected (i.m.) with saline or 200 mug of GnRH at 33 to 35 days postpartum. Saline or GnRH was injected 5 hr before calf return. Plasma luteinizing hormone (LH) was measured in blood samples collected every 30 min for 5.5 hr beginning 30 min prior to injection of saline or GnRH. Plasma progesterone was measured in blood samples collected 0, 7, and 14 days after GnRH injection and 7 and 14 days following the first detected estrus. There were no differences (P>0.05) in the interval to peak LH release or the magnitude of the LH release between the GnRH and CR plus GnRH groups; however, the GnRH induced release of LH was greater (P<0.05) over time when preceded by CR. Plasma progesterone concentrations were increased on day 7, compared to day 0, after GnRH injection in 57% and 50% of the animals in the GnRH and CR plus GnRH groups, respectively. However, behavioral estrus was not observed in any of the cows between days 0 and 7 after GnRH injection. A higher (P<0.05) percentage of the cows injected with GnRH formed luteal tissue compared to cows injected with saline; however, the luteal lifespan following GnRH injection was decreased relative to the luteal lifespan following the first observed estrus. The mean interval from calving to first estrus was decreased (P<0.05) by 17 days in the CR group relative to the other groups, and calf removal had no detrimental effect on milk production at 80 days postpartum or on calf weaning weights at approximately 7 months of age. In summary, 72 hr CR decreased the postpartum interval and increased the pituitary responsiveness to GnRH. Pretreatment with 72 hr CR did not alter circulating progesterone concentrations or luteal lifespan of corpora lutea induced by GnRH.     

Activation and nuclear translocation of PKCdelta,Pyk2 and ERK1/2 by gonadotropin releasing hormone in HEK293 cells

The mechanism of agonist-induced activation of Pyk2 and its relationship with ERK1/2 phosphorylation was analyzed in HEK293 cells stably expressing the gonadotropin releasing hormone (GnRH) receptor. GnRH stimulation caused rapid and sustained phosphorylation of ERK1/2 and Pyk2 that was accompanied by their nuclear translocation. Pyk2 was also localized on cell membranes and at focal adhesions. Dominant negative Pyk2 (PKM) had no effect on GnRH-induced ERK1/2 phosphorylation and c-fos expression. These actions of GnRH on ERK1/2 and Pyk2 were mimicked by activation of protein kinase C (PKC) and were abolished by its inhibition. GnRH caused translocation of PKC and δ, but not of , ι and λ, to the cell membrane, as well as phosphorylation of Raf at Ser338, a major site in the activation of MEK/ERK1/2. Stimulation of HEK293 cells by EGF caused marked ERK1/2 phosphorylation that was attenuated by the selective EGFR receptor (EGF-R) kinase inhibitor, AG1478. However, GnRH-induced ERK1/2 activation was independent of EGF-R activation. These results indicate that activation of PKC is responsible for GnRH-induced phosphorylation of both ERK1/2 and Pyk2, and that Pyk2 activation does not contribute to GnRH signaling. Moreover, GnRH-induced phosphorylation of ERK1/2 and expression of c-fos in HEK293 cells is independent of Src and EGF-R transactivation, and is mediated through the PKC/Raf/MEK cascade.     

Correlation of seasonal changes in sperm output with endocrinological changes in the adult male bonnet monkey,Macaca radiata

We have examined the monthly variations in sperm output and attempted to correlate the profiles of endocrine hormones secreted with the sperm counts throughout the year in the adult male bonnet monkey. As previously reported, there was a distinct spurt in sperm output beginning September through December months. A concomitant increase in serum testosterone and prolactin concentrations were also noted during September through November (mid and post-monsoon season). Although there was a marked increase in gonadotropin releasing hormone stimulated testosterone secretion, the peak testosterone concentrations post gonadotropin releasing hormone injection did not vary significantly (P > 0.05) throughout the year. Basal serum follicle stimulating hormone concentrations did not vary significantly (P > 0.05) during April to June months compared to September-November months. Serum inhibin concentration remained unaltered throughout the year, except in the month of March. The results of this study provide evidence for annual rhythms in prolactin and testosterone secretion and a distinct seasonality in the sperm output of the adult male bonnet monkey, but the pituitary responsiveness to exogenous gonadotropin releasing hormone remains unaltered throughout the year. Because of the existence of seasonality as noted in the present study, future studies which utilize the adult male bonnet monkey as an experimental model need to take into consideration the seasonal effects on reproductive function in this species.     

Signal transduction of the gonadotropin releasing hormone (GnRH) receptor: Cross-talk of calcium,protein kinase C (PKC), and arachidonic acid

Summary 1. The decapeptide neurohormone gonadotropin releasing hormone (GnRH) is the first key hormone of the reproductive system. Produced in the hypothalamus, GnRH is released in a pulsatile manner into the hypophysial portal system to reach the anterior pituitary and stimulates the release and synthesis of the gonadotropin hormones LH and FSH. GnRH, a Ca²⁺ mobilizing ligand, binds to its respective binding protein, which is a member of the seven transmembrane domain receptor family and activates a G-protein (Gq).2. The subunit of Gq triggers enhanced phosphoinositide turnover and the elevation of multiple second messengers required for gonadotropin release and biosynthesis.3. The messenger molecules IP₃, diacylglycerol, Ca²⁺, protein kinase C, arachidonic acid and leukotriene C₄ cross-talk in a complex networks of signaling, culminating in gonadotropin release and gene expression.     

Annual cycle of activity of the neurons of the anterior preoptic area in the brain of Rana esculenta L.

Abstract

Sexually mature male and female Rana esculenta L. were captured in their natural habitat in six phases of the annual cycle. Nuclear volumes in APOA cells were found to fluctuate distinctly in the course of the year. In both sexes nuclear volumes were maximal in the phases preceding the breeding season (IIIrd decade of January, and 1st decade of April), and minimal throughout the phases of active life (IIIrd decade of May, IInd decade of July, and 1st decade of September). No aldehydefuchsin or Gomori‐positive material was found in the APOA perikaryons.     

Synaptology of luteinizing hormone-releasing hormone neurons in rat preoptic area

The ultrastructural appearance of luteinizing hormone-releasing hormone (LHRH) immunoreactive elements was studied in the medial preoptic area (MPOA) of adult male Fischer 344 rats. The purpose of the study was to determine the distribution and morphology of innervation of the LHRH neuron. Although not numerous, both axo-somatic and axo-dendritic synapses were present and generally of the asymmetric (Gray's II) category. Analyses of 56 profiles of 11 separate perikarya revealed only 7 axo-somatic terminals. The synaptic input to LHRH dendrites was a fraction of that to non-identified dendrites in the same electron micrographic fields; 0.4% of LHRH dendritic membrane was in synaptic contact compared to 6.6% of nonidentified dendritic membrane. In addition to receiving an input, LHRH processes were also seen to make synapses onto non-immunoreactive elements. Close examination of this material for evidence of contact between LHRH elements revealed two clear examples of synaptic interaction and several instances of close association in which no other elements intervened.     

Localization of putative gonadotrophin releasing hormone receptor protein in the anterior pituitary

Specific binding of a fully biologically active 125I-gonadotrophin releasing hormone (GnRH) to isolated anterior pituitary cells is time dependent, saturable and the concentration dependent binding curves exhibit positive cooperativity. Binding to intact or solubilized plasma membranes and an affinity purified GnRH receptor protein reveals in all instances multiple high affinity binding sites. Thus, GnRH receptor protein appears to be an intrinsic constituent of the cell membrane, and perhaps, other membranous organelles. To investigate the latter, the binding of 125I-GnRH to various subcellular fractions was studied and its affinity and time requirements determined. GnRH binding to plasma membranes and secretory granules was to multiple high affinity sites, while that to nuclei and microsomes was to a single high affinity site. Binding was 1.83 +/- 0.07, 0.78 +/- 0.04, 0.31 +/- 0.03 and 0.27 +/- 0.03 fmol micrograms-1 protein for isolated plasma membranes, secretory granules, microsomes and nuclei, respectively, after 30 min incubation with 10(-9) M GnRH. The magnitude of binding to microsomes did not change during the incubation period. It did not show any decrease (p greater than 0.05) in isolated nuclei and plasma membranes, except for the 24 h time period, when a significant drop (p less than 0.001) was seen. Binding to the secretory granule fraction culminated at 15 min and then decreased (p less than 0.001) steadily to a non-detectable level at 24 h. Thus GnRH receptor protein or its portion may be an integral part of some membranous particles in the anterior pituitary cells.(ABSTRACT TRUNCATED AT 250 WORDS)