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Biomedical subjects

A J Silverman

Publications and source records attributed to A J Silverman.

At least 73 records · Page 4Linked to original sources
PubMed1990

Repair of reproductive deficits by neural transplantation.

Transplantation of brain tissue has been used to ameliorate the genetic lesion of the hypogonadal mutant mouse. This animal does not synthesize gonadotropin-releasing hormone (GnRH) and so has an infantile reproductive system. Implantation of normal fetal or neonatal preoptic area containing GnRH neurons reverses many aspects of the reproductive deficiency. Pituitary and plasma levels of gonadotropins rise, followed by growth of the gonads and sexual organs. Pituitary release of gonadotropins is episodic, suggesting that the grafted tissue is integrated into the "pulse generator." The vast majority of grafted animals do not show castration-induced elevations of luteinizing hormone (LH) nor respond to exogenous steroids with a depression in circulating LH. Negative feedback of gonadal steroids seems to be inoperative. In contrast, some females can show ovulatory surges of LH in response to mating (reflex ovulation), after administration of exogenous steroid (progesterone), and, on rare occasion, ovulation cycles occur spontaneously. Anatomical studies demonstrate that reproductive recovery is dependent on the outgrowth of GnRH axons to the host median eminence. Some but not all of the GnRH neurons within the grafts contribute to this innervation. GnRH axons exit into the host along well-defined pathways, recapitulating in part the paths taken by normal axons. How the graft and host are integrated to produce the panoply of reproductive responses is the subject of current study.

Animals↗
PubMed1990

Light and electron microscopic immunocytochemical analysis of antibodies directed against GnRH and its precursor in hypothalamic neurons.

A battery of antibodies directed against different portions of the precursor to gonadotropin-releasing hormone (GnRH), as well as to the mature decapeptide, were characterized immunocytochemically in two ways. Absorption experiments were used to determine the epitope recognized by each antiserum. Electron microscopic immunocytochemistry was then used to define the subcellular organelles that contained reaction product when tissue was incubated with these reagents. These latter observations helped to determine if the antibody recognized the epitope as part of the intact precursor or only after it had been cleaved from parent protein. Our results demonstrate that the GnRH precursor is routed from the rough endoplasmic reticulum through the Golgi apparatus to the secretory vesicles. Furthermore, we show that initial cleavage and processing of the GnRH precursor begin in the cell soma. These antibodies should be useful in the future in determining changes in processing of precursor in animals that differ in endocrine function.

Amino Acid Sequence↗
PubMed1990

Sexual dimorphism in the synaptic input to gonadotropin releasing hormone neurons.

GnRH neurons form the final common pathway regulating the secretion of gonadotropins from the anterior pituitary. Since the patterns of gonadotropin release display profound sexual dimorphism among mammals including the rodent, we undertook an ultrastructural analysis to determine whether these neurosecretory cells were differentially innervated between the sexes. As a further exploration of the organization of the neurocircuitry integrating GnRH neurons with the central nervous system, we also determined the degree to which GnRH cells and their processes were innervated by terminals containing either the endogenous opiate, beta-endorphin (BE) or GnRH itself. Sections from the diagonal band of Broca and the preoptic area of adult male and diestrus II female rats were immunocytochemically processed for dual localization of GnRH and BE. GnRH neurons cut through the plane of the nucleus were identified in 1 micron sections. Serial ultrathin sections were made and analyzed for 1) total synaptic input to both cell bodies and dendrites; 2) BE input; and 3) input arising from GnRH itself. We report that GnRH neuronal cell bodies in females received approximately twice the number of synapses as did those of males. The input to the GnRH dendrites, when measured as percent of plasma membrane in synaptic contact, also showed a profound sexual dimorphism with the female having a larger proportion of the dendrite in synaptic apposition. BE innervation contributed to this dimorphism at the level of both the cell body and dendrite. In contrast, the distribution and number of GnRH terminals did not differ between the sexes. In both they were confined to the dendritic arbor. We hypothesize that the capacity of the female rodent GnRH system to show neurogenic derived alterations in GnRH output not seen in the male may be due in part to these anatomical differences.

Animals↗
PubMed1989

Identification of gonadotropin releasing hormone (GnRH) neurons projecting to the median eminence from third ventricular preoptic area grafts in hypogonadal mice.

Functional gonadotropin releasing hormone (GnRH) neurosecretory activity can be restored in genetically hypogonadal (hpg) adult mice with grafts of GnRH-containing fetal or neonatal septal-preoptic area (S/POA) tissue. Neurons implanted into the third ventricle of the host brain survive and send out axons which innervate one of the normal targets of these neurons, the median eminence (ME). Fibers terminate near primary portal vessels where GnRH is available for release into the vasculature, and this axonal outgrowth is essential for the stimulation of gonadotropin secretion, gonadal and accessory sex structure growth, gametogenesis, and fertility. Although it is known that GnRH axons reach their target, it is not known if all such neurons in a graft contribute to the projection. Taking advantage of the fact that axons in the ME, the sole host target, are outside the blood-brain barrier (BBB), long-term grafted animals were injected intraperitoneally with a retrograde tracer, Fluorogold (FG). Normal male mice were injected for comparison. Animals were sacrificed 5 days after injection and brain sections in the area of the graft were stained immunocytochemically for GnRH. In the normal male mice, two-thirds of the GnRH neurons were double-labeled with FG. In grafted individuals which showed increased gonadal growth, the percentage of labeled cells ranged from 17 to 75%. The results indicate that despite tissue injury, ectopic location, and a vastly reduced population, transplanted fetal GnRH neurons recapitulate a pattern seen in normal intact mice where some but not all neurons were capable of capturing a peripherally delivered tracer.

Animals↗
PubMed1989

beta-Endorphin and gonadotropin-releasing hormone synaptic input to gonadotropin-releasing hormone neurosecretory cells in the male rat.

Physiological and pharmacological evidence has suggested that both endogenous opiates and gonadotropin-releasing hormone (GnRH) itself can act centrally to exert a tonic inhibition on gonadotropin secretion via an inhibition of the neurosecretion of GnRH. To determine if the effects of these two peptides might be mediated via a direct synaptic input to the GnRH neuron, we undertook a double label ultrastructural study. We were able to localize in the same tissue section beta-endorphin and GnRH. Analysis of serial sections through GnRH perikarya and dendrites in the male rat diagonal band/preoptic area revealed that almost 10% of the synapses impinging on the GnRH neuron contained beta-endorphin; an additional 10% of the terminals contained GnRH. These data provide anatomical evidence in support of both a direct modulation of GnRH release by opiates and of the presence of an ultrashort feedback loop.

Animals↗
PubMed1989

Estradiol-concentrating cells in the brains of hypogonadal female mice and in their intraventricular preoptic area implants.

Estradiol-concentrating cells were evaluated in the brains of hypogonadal female mice and in their intraventricular preoptic area brain grafts using autoradiography for [3H]estradiol. Normal distribution of estradiol-concentrating cells was observed in the brains of the hypogonadal mice with dense collections of these cells in the lateral septum; the medial preoptic area; the medial anterior hypothalamus; the ventromedial, arcuate, and periventricular nuclei of the hypothalamus; and the medial and cortical nuclei of the amygdala. In addition, estradiol-concentrating cells were present in all the transplants, with the estimated number of such cells in the transplants ranging from 390 to 2600. There was no correlation between numbers of estradiol-concentrating cells within the transplants and degree of reproductive recovery in the hypogonadal mice. Gonadotropin-releasing hormone (GnRH) immunocytochemistry of alternate sections revealed GnRH-immunoreactive material within the grafts and immunoreactive fibers exiting the grafts and entering the hosts' median eminence. No specific relationship between GnRH cells and estradiol-concentrating cells was evident within the grafts, nor was there any indication of identity of estrogen-concentrating cells with GnRH cells.

Animals↗
PubMed1989

Corticotropin-releasing factor synapses within the paraventricular nucleus of the hypothalamus.

Corticotropin-releasing factor (CRF) regulates the release of adrenocorticotropin (ACTH) from the anterior pituitary and these neurosecretory neurons reside in the paraventricular nucleus of the hypothalamus (PVN). In addition to its role as an ACTH secretogogue, exogenously administered CRF can act centrally to modify sympathetic outflow, alter various stress-induced behaviors and modulate its own secretion. Some of these effects might be mediated by CRF acting synaptically within the PVN as the nucleus is known to play a major role in integration of autonomic function. The current ultrastructural immunocytochemical study was designed to examine the range of synaptic relationships that CRF terminals make within the PVN. CRF-positive synapses were numerous, particularly in the periventricular zone. The majority of terminals formed axo-dendritic synapses and of these over 85% were the Gray's type II (symmetrical) class. Axo-somatic terminals were also encountered and both parvicellular and magnocellular neurons were innervated. Once again most of the terminals were Gray's type II. Although an innervation of unidentified structures was the most common, CRF synapses onto CRF neurons and dendrites were observed. All CRF/CRF interactions had symmetrical membrane specializations. These studies indicate that CRF could play a prominent role in the modulation of both parvicellular and magnocellular neurons within the paraventricular nucleus, including modulation of its own neurosecretory activity.

Animals↗
PubMed1989

Effects of gonadectomy and treatment with gonadal steroids and luteinizing hormone secretion in hypogonadal male and female mice with preoptic area implants.

The ability of male and female hypogonadal (hpg) mice with preoptic area (POA) grafts to show negative feedback was studied. GnRH neurons within POA grafts send axons into the median eminence of the hpg host (HPG/POA), resulting in increased gonadotropin production and gonadal development in the mice that are genetically unable to produce GnRH. The present studies evaluated whether negative feedback, an aspect of normal reproductive function, is present in male and female HPG/POA mice. In normal male mice plasma LH was increased 1 or 5 months after castration and returned to baseline after testosterone propionate treatment. In contrast, no alterations in plasma LH were measured in similarly treated HPG/POA males. HPG/POA female mice were ovariectomized 6 weeks or 3 or 6 months after graft surgery and received sc 17 beta-estradiol (E2) implants 3 months later. Normal mice were studied when 6 weeks old and 8 months old (age-matched to HPG/POA mice ovariectomized 3 months after graft surgery). Further, to determine whether the mice were capable of positive feedback, 1 week after receiving E2 implants, mice in the 6-week and 3-month postgraft surgery groups were challenged with sequential administration of estradiol benzoate and progesterone. The significant increase in plasma LH after ovariectomy or decrease after E2 implant in normal female mice was not present in most HPG/POA female mice. Just 2 of the 24 HPG/POA females studied had increased plasma LH after gonadectomy, and in only 1 of these was plasma LH suppressed by E2 treatment. The ability of an individual HPG/POA mouse to show positive feedback did not predict the ability to show negative feedback, nor did the ability to show negative feedback predict positive feedback capability. Among the mice that failed to respond to ovariectomy with increased LH release were some that had elevated LH in response to steroid challenge or had spontaneously ovulated. On the other hand, neither mouse that had increased LH release after ovariectomy had shown positive feedback to a steroid challenge. Immunocytochemical evaluation revealed GnRH cells within the grafts and GnRH fiber innervation of the host's median eminence, but there was no correlation between numbers of GnRH cells or extent of innervation with the ability to show either negative or positive feedback, nor was the presence of vasoactive intestinal peptide cells within the grafts of predictive value. The failure of negative feedback in most of the HPG/POA mice tested may be due to the failure to establish as yet unidentified but essential afferents to the grafted GnRH cells and/or their axonal processes.

Animals↗
PubMed1988

Ultrastructure and synaptic organization of luteinizing hormone-releasing hormone (LHRH) neurons in the anestrous ewe.

Electron microscopic immunocytochemistry was employed to examine the ultrastructure of luteinizing hormone-releasing hormone (LHRH) neurons and their projections to the median eminence in the sheep brain. LHRH perikarya in the preoptic area of anestrous ewes are less innervated than nonimmunoreactive cells in the same sections, but still receive numerous synaptic inputs, primarily onto distal dendrites and small somatic protuberances. Axon terminals synapsing upon LHRH cells contain a combination of clear spherical vesicles and larger dense-core vesicles. Interestingly, LHRH cell bodies and dendrites are almost entirely surrounded by glial processes. These processes intervene between immunoreactive elements that at a light microscopic level appear to be in contact with each other. Thus no evidence was obtained at the ultrastructural level for contacts among adjacent LHRH cells or dendrites in the preoptic area. Synaptic inputs onto LHRH cell bodies and dendrites appear to penetrate this glial sheath. In contrast to the absence of contacts among LHRH cells in the preoptic area, individual LHRH terminals in the median eminence are often clustered in direct plasma membrane contact. Comparisons between animals of differing reproductive status are needed to determine whether alterations in synaptic inputs, glial ensheathment, or LHRH-LHRH appositions, may underlie seasonal changes in the activity of LHRH neurons.

Anestrus↗
PubMed1988

Quantitative analysis of synaptic input to gonadotropin-releasing hormone neurons in normal mice and hpg mice with preoptic area grafts.

Mutant hypogonadal (hpg) mice with a truncated gene for the precursor to gonadotropin-releasing hormone (GnRH) show certain aspects of recovery of reproductive function after receiving grafts of normal preoptic area into the third ventricle. We have previously shown that GnRH neurons from within the grafts can innervate the appropriate neural-hemal target in the host. To determine if in turn these exogenously derived neurons receive a synaptic input comparable to the GnRH neurons in the normal animal we have now carried out a quantitative ultrastructural analysis to compare the synaptic input to GnRH neurons in the normal preoptic area and in the grafts. In almost all cases GnRH cells or dendrites in normal brains and within the grafts received a synaptic input. In normal animals, input to GnRH dendritic profiles was significantly greater (P less than 0.001) than to the somatic plasma membrane and this trend was also observed within the grafts though the difference was not statistically significant. In addition, no statistically significant difference was found between the input to GnRH structures within the grafts and in normal preoptic area. However, a substantial variability in input among grafted animals was evident which was not observed in normal animals. The sources of variability within the grafts are discussed and we suggest that the deficiencies and differences that exist in regulation of gonadotropin secretion among grafted hpg animals may be reflected in aberrant synaptic input.

Animals↗
PubMed1988

Potential sites of interaction between catecholamines and LHRH in the sheep brain.

A combined immunoperoxidase/immunofluorescence procedure was used to examine potential sites of overlap between catecholamine and LHRH systems in the brains of ewes sacrificed during either anestrous or the breeding season. Cells and fibers immunoreactive for either tyrosine hydroxylase (TH) or dopamine-beta-hydroxylase (DBH) were visualized in the same sections as immunopositive LHRH perikarya and fibers. TH- and DBH-positive varicosities in the preoptic area and anterior hypothalamus appeared to contact both LHRH cell bodies and their dendrites. Clusters of TH-positive cells and fibers were found in the organum vasculosum of the lamina terminalis, and partially overlapped the location of immunoreactive LHRH fibers in that structure. Immunoreactive TH and LHRH fibers were densely interspersed within the zona externa of the median eminence, particularly within its lateral portion. No obvious qualitative differences were apparent in either the distribution of catecholamine cells and fibers or their overlap with LHRH elements between the brains of anestrous and breeding season ewes. These observations suggest the possibility of catecholaminergic synaptic inputs onto LHRH neurons in the ewe, as well as the potential for interaction between catecholamines and LHRH at the level of the median eminence.

Animals↗
PubMed1988

Ultrastructure of luteinizing hormone-releasing hormone (LHRH) neurons and their projections in the golden hamster.

Seasonal breeding mammals exhibit a reversible annual cycle of fertility, and thus represent valuable models for investigating the organization of luteinizing hormone-releasing hormone (LHRH) neurons which mediate the neuroendocrine control of reproduction. Electron microscopic immunocytochemistry was used to examine the ultrastructure of LHRH neurons and their projections in a seasonal breeder, the golden hamster, housed under photoperiodic conditions which are reproductively stimulatory. LHRH perikarya in the diagonal band, medial septum, and preoptic area were bipolar or unipolar cells which possessed nuclei with prominent indentations and multiple nucleoli. LHRH reaction product within these cells was associated with neurosecretory granules and the rough endoplasmic reticulum (RER), particularly those portions of RER adjacent to the outer nuclear envelope. LHRH cell bodies and dendrites in the hamster received an extremely limited amount of synaptic input; of a total of twenty-five cells analyzed, we found only five instances of morphologically identified synapses onto immunoreactive dendrites. Occasionally close somatic appositions were seen between LHRH cells and non-immunoreactive neurons, and in one instance between two LHRH somas. In the organum vasculosum of the lamina terminalis (OVLT), LHRH fibers appeared as isolated strings of varicosities. In contrast, within the median eminence immunoreactive axonal profiles were frequently bundled together in fascicles although synaptic specializations were not observed between individual axons. The existence of contacts between LHRH axons in the median eminence, coupled with the extreme paucity of synaptic inputs onto LHRH neurons in this species, suggests that the median eminence may be a site where neural or hormonal signals could influence the coordinated release of LHRH from cells of dispersed origin.

Animals↗
PubMed1988

Transplanted gonadotropin-releasing hormone neurons promote pulsatile luteinizing hormone secretion in congenitally hypogonadal (hpg) male mice.

Congenitally hypogonadal (hpg) male mice are unable to synthesize biologically active gonadotropin-releasing hormone (GnRH). Implantation of normal fetal preoptic area tissue containing GnRH neurons into the third ventricle of adult hpg males significantly elevates pituitary levels of luteinizing hormone (LH) and corrects their hypogonadism. In all responding animals, immunoreactive GnRH neurons within the transplant innervate the median eminence of the host. To assess whether gonadal recovery in hpg hosts results from pulsatile secretion of GnRH from grafted neurons, we compared the pattern of variation in plasma LH levels in 19 hpg graft recipients with testicular growth to that of 10 normal adult mice. All animals were castrated prior to receiving an indwelling catheter in the jugular vein. Sequential blood samples were collected (t = 10 min) and assayed for LH. Pulsatile LH secretion was seen in 11 of 19 hpg hosts and in all control mice. While there was great variability between individual animals, measures of baseline LH, LH pulse amplitude and duration, interpulse interval, and LH pulse frequency revealed no difference between hpg graft recipients and normal castrates in their LH pulse pattern. Immunocytochemical analysis of the brain in hpg hosts suggested no correlation between any parameter of pulse activity and individual differences in GnRH cell number or GnRH fiber outgrowth into the median eminence. Sources of variation in LH secretion among graft recipients, and between hpg hosts and normal mice, are discussed. We suggest that transplanted GnRH neurons are capable of integration into a GnRH 'pulse generator' which can support a near-normal pattern of pulsatile LH secretion, leading to testicular growth and steroid production.

Animals↗
PubMed1988

Positive feedback in hypogonadal female mice with preoptic area brain transplants.

When fetal preoptic area (POA) brain grafts that contain gonadotropin-releasing hormone cells are transplanted into the third ventricle of adult female hypogonadal mice, the animals respond with sexual maturation, persistent estrus, and the ability to ovulate reflexively after mating. However, the absence of normal spontaneous ovulatory cyclicity suggests an impairment in positive feedback. We, therefore, studied the effect of administration of progesterone alone or of sequential estradiol benzoate and progesterone on plasma levels of luteinizing hormone (LH) in groups of hypogonadal (HPG) mice in persistent estrus after receiving POA grafts (HPG/POA). Individual differences in responsivity to progesterone were related in part to the length of time in persistent estrus. Approximately 30% of HPG/POA grafts tested 2 months after graft showed increased levels of plasma LH. This was reduced to 10% when animals were tested 5 months after graft. Sequential administration of estradiol benzoate plus progesterone to intact HPG/POA mice was ineffective in elevating LH. The presence of corpora lutea in ovaries verified that only animals with a progesterone induced LH surge ovulated. Other HPG/POA mice were mated, and the occurrence of reflex ovulation was determined. Four of these mice delivered pups: 3 were previous responders to progesterone. One female mated again during the immediate postpartum period and delivered a second litter. Following weaning of all offspring, this animal displayed spontaneous ovarian cyclicity, confirmed by ovarian histology. This is the first proven example of spontaneous ovulation in a mutant mouse with a brain graft. The results show that some HPB/POA mice are capable of positive feedback responses, and rarely, of becoming spontaneous ovulators.

Animals↗
PubMed1987

Plasma LH rises rapidly following mating in hypogonadal female mice with preoptic area (POA) brain grafts.

Hypogonadal female mice, genetically deficient in gonadotropin releasing hormone (GnRH), respond to preoptic area (POA) grafts obtained from normal fetal or neonatal mice with increased gonadotropin levels, ovarian and uterine development and continual vaginal estrus rather than spontaneous ovulatory cyclicity. Previous studies showed that such mice became pregnant following one overnight pairing with a normal male, indicating reflex ovulation. The present study evaluated plasma LH concentrations in relation to mating. Plasma LH levels in the hpg females with POA grafts were significantly elevated 10 min following the male partner's ejaculation, but were no different than baseline at 30, 60 or 120 min following the male's ejaculation. The post-copulatory plasma LH levels of 3.0 +/- 0.6 ng/ml (mean +/- S.E.M.) were considerably lower than the proestrous LH surge seen in the normal females in the colony (16.8 +/- 4.8 ng/ml), but in at least 4 of 10 hpg mice the levels were sufficient to induce ovulation as proved by pregnancy following this single mating. Grafts contained GnRH-reactive cells and fibers that projected to the median eminence of the host brains.

Animals↗
PubMed1987

Accessory olfactory bulb transplants correct hypogonadism in mutant mice.

Transplantation of normal fetal gonadotropin-releasing hormone (GnRH) neurons from the accessory olfactory bulb (AOB) to the third ventricle of GnRH-deficient adult mutant mice reverses the genetically determined reduction in pituitary hormones and poorly developed gonads. The transplanted heterotopic AOB neurons adapt their morphology and secretory functions to what is observed with preoptic GnRH neurons when transplanted into deficient mice and in the normal intact mature animal. This suggests the presence of median eminence trophic factors affecting the growth, terminal sprouting, and functional behavior of the transplanted neurons.

Animals↗