A Study of the Gonadotropin Releasing Hormone Neuronal Network in the Median Eminence of the Rhesus Monkey ( Macaca mulatta) Using a Post-Embedding Immunolabelling Procedure

The purpose of this study was to describe the ultrastructural features of gonadotropin releasing hormone (GnRH) axonal processes in the median eminence of the monkey, using a post-embedding immunogold labelling procedure. Evidence was also sought to evaluate the view that release of this peptide may be governed by direct inputs to GnRH axons in the median eminence. Plastic embedding was used to preserve ultrastructure, and a polyclonal rabbit anti-GnRH was used as primary antibody. Immunogold labelling with 15-nm particles was almost exclusively found overlying dense core vesicles (dcvs) and preabsorption of the primary antibody with synthetic GnRH eliminated this labelling. Morphometric analysis was performed on tissue from two monkeys. Four types of profiles containing GnRH immunoactive dcvs were observed. Type I profiles were morphologically unremarkable with a cross sectional area of approximately 0.6 microm2 and probably represent intervaricose axon segments. Type II profiles, which were nominally larger than Type I structures, were characterized by a high density of round microvesicles, which were frequently concentrated along the neuronal membrane to form 'synaptoid' contacts with adjacent glia. Two additional and large GnRH profiles (>5 microm2) were observed. One (Type III) contained a high density of dcvs and mitochondria, and was considered analogous to an axonal swelling or Herring body in the magnocellular hypothalamo-neurohypophysial system. The Type IV structure, which was considered not to be a Herring body because of the relative low density of mitochondria was innervated by a classical symmetrical synapse. The functional significance of these observations is discussed.     

The gonadotropin-releasing hormone containing ventral hypothalamic tract in the fetal rhesus monkey (Macaca mulatta)

A well-defined, gonadotropin-releasing hormone (GnRH)-containing fiber pathway, the ventral hypothalamic tract (VHT), is described by immunostaining in fetal rhesus macaques (109-156 days gestation). The VHT arises above the lateral aspects of the optic chiasm near the supraoptic nucleus, and courses ventromedially close to the ventral hypothalamic surface to terminate in the infundibulum and zona externa of the median eminence. It is formed by the confluence of GnRH-immunopositive (GnRH+) axons from local neurons, from a few GnRH+ cells in the inferior thalamic peduncle, and probably from more anterior neurons in the septum and preoptic area. Bipolar GnRH+ neurons contributing directly to the VHT are grouped at its origin dorsolateral to the optic chiasm, dorsal and medial to the optic tracts, at the infundibular lip, and within the pathway between. At the infundibular lip, GnRH+ perikarya are generally lateral or ventral to the infundibular (arcuate) nucleus, and are rarely within the nucleus itself. Cell bodies here are sometimes tripolar, but GnRH+ intercellular contacts are seldom seen. A few VHT fibers extend to the ventral surface of the brain just beneath the pia mater. Abundant capillaries in the subarachnoid space suggest a possible route for delivery of GnRH to the adenohypophysis in early gestation, before maturation of the hypophysial portal system occurs. Posterior to the infundibulum, a few VHT fibers are joined by descending periventricular fibers forming a dense fiber band beneath the premammillary recess of the third ventricle. Totals of GnRH+ cell bodies in the prosencephalon of the fetal rhesus macaque are estimated to be 5,600 in females (n = 2) and 2,600 in males (n = 3). More than 60% of VHT neurons are located in the medial basal hypothalamus, and the majority of basal hypothalamic GnRH+ neurons (86%) are associated with the VHT. Furthermore, reports of the autonomy of the medial basal hypothalamic-hypophysial unit in control of gonadotropin secretion suggest that the VHT may be the most important GnRH system involved in primate reproduction. It is clear that fetal material may offer the best model to study the GnRH neuronal system in primates.     

Gonadotrophin‐Releasing Hormone (GnRH) Exerts Stimulatory Effects on GnRH Neurones in Intact Adult Male and Female Mice

There is substantial evidence for a role of the neuropeptide gonadotrophin‐releasing hormone (GnRH) in the regulation of GnRH neurone secretion but how this is achieved is not understood. We examined here the effects of GnRH on the electrical excitability and intracellular calcium concentration ([Ca²⁺]_i) of GnRH neurones in intact adult male and female mice. Perforated‐patch electrophysiological recordings from GnRH‐green fluorescent protein‐tagged GnRH neurones revealed that 3 nm –3 μm GnRH evoked gradual approximately 3 mV depolarisations in membrane potential from up to 50% of GnRH neurones in male and female mice. The depolarising effect of GnRH was observed on approximately 50% of GnRH neurones throughout the oestrous cycle. However, at pro‐oestrus alone, GnRH was also found to transiently hyperpolarise approximately 30% of GnRH neurones. Both hyperpolarising and depolarising responses were maintained in the presence of tetrodotoxin. Calcium imaging studies undertaken in transgenic GnRH‐pericam mice showed that GnRH suppressed [Ca²⁺]_i in approximately 50% of GnRH neurones in dioestrous and oestrous mice. At pro‐oestrus, 25% of GnRH neurones exhibited a suppressive [Ca²⁺]_i response to GnRH, whereas 17% were stimulated. These results demonstrate that nm to μm concentrations of GnRH exert depolarising actions on approximately 50% of GnRH neurones in males and females throughout the oestrous cycle. This is associated with a reduction in [Ca²⁺]_i. At pro‐oestrus, however, a further population of GnRH neurones exhibit a hyperpolarising response to GnRH. Taken together, these studies indicate that GnRH acts predominantly as a neuromodulator at the level of the GnRH cell bodies to exert a predominant excitatory influence upon GnRH neurones in intact adult male and female mice.     

Morphology of the inferior olivary complex of the rhesus monkey (Macaca mulatta)

The morphology of the inferior olivary complex was determined in the rhesus monkey (Macaca mulatta). The position, configuration, and relations of each of its components were ascertained in serial transverse sections extending from caudal through rostral poles of the complex. The medial accessory nucleus was divided into seven cell groups (labeled a through g). The principal nucleus consisted of well-developed dorsal, ventral, and lateral lamellae and a criterion was established for distinguishing the boundaries of each. This distinction was based upon the pattern of invagination that developed in the principal nucleus in progressing rostrally from its caudal pole. The dorsal and lateral lamellae each presented a characteristic pattern of major invaginations, while the ventral lamella did not. Finally, a detailed comparison was made of the inferior olivary complex of the monkey with that of the cat as determined by other investigators. It was shown that the medial and dorsal accessory nuclei of the two species appeared similar and that the major difference between these species concerns the principal nucleus.     

Ovarian Regulation of Kisspeptin Neurones in the Arcuate Nucleus of the Rhesus Monkey (Macaca mulatta)

Tonic gonadotrophin secretion throughout the menstrual cycle is regulated by the negative‐feedback actions of ovarian oestradiol (E₂) and progesterone. Although kisspeptin neurones in the arcuate nucleus (ARC) of the hypothalamus appear to play a major role in mediating these feedback actions of the steroids in nonprimate species, this issue has been less well studied in the monkey. In the present study, we used immunohistochemistry and in situ hybridisation to examine kisspeptin and KISS1 expression, respectively, in the mediobasal hypothalamus (MBH) of adult ovariectomised (OVX) rhesus monkeys. We also examined kisspeptin expression in the MBH of ovarian intact females, and the effect of E₂, progesterone and E₂ + progesterone replacement on KISS1 expression in OVX animals. Kisspeptin or KISS1 expressing neurones and pronounced kisspeptin fibres were readily identified throughout the ARC of ovariectomised monkeys but, on the other hand, in intact animals, kisspeptin cell bodies were small in size and number and only fine fibres were observed. Replacement of OVX monkeys with physiological levels of E₂, either alone or with luteal phase levels of progesterone, abolished KISS1 expression in the ARC. Interestingly, progesterone replacement alone for 14 days also resulted in a significant down‐regulation of KISS1 expression. These findings support the view that, in primates, as in rodents and sheep, kisspeptin signalling in ARC neurones appears to play an important role in mediating the negative‐feedback action of E₂ on gonadotrophin secretion, and also indicate the need to study further their regulation by progesterone.     

去卵巢及去松果体对大鼠纹状体细胞凋亡的影响

目的　探讨去卵巢 (OVX)、去松果体 (PX)对纹状体细胞凋亡的影响。方法　将大鼠随机分为正常、假OVX、OVX、假 PX、PX、假 OVX+假 PX、OVX+PX7组。术后 90 d,用 HE、TUNEL染色观察 OVX、PX对纹状体细胞凋亡的影响。结果　假手术各组与正常组在各项检测指标上无明显差异 ,合并为 A组 ;A组在纹状体偶有细胞凋亡但部位不恒定。与 A组比 ,OVX、PX使纹状体细胞凋亡数显著增多 (P <0 .0 1 ) ,并且 PX+OVX >PX>OVX(P <0 .0 1 ) ;各组纹状体无明显胶质细胞增生 (P >0 .0 5 )。结论　 OVX、PX可诱发纹状体细胞凋亡 ,而且 PX比 OVX影响大 ,OVX+PX影响更大 ;这种影响与胶质细胞无关。     

Localization of nAChR subunit mRNAs in the brain of Macaca mulatta

We present here a systematic mapping of nAChR subunit mRNAs in Macaca mulatta brain. A fragment, from the transmembrane segments MIII to MIV of Macaca neuronal nAChR subunits was cloned, and shown to exhibit high identity (around 95%) to the corresponding human subunits. Then, specific oligodeoxynucleotides were synthesized for in situ hybridization experiments. Both alpha4 and beta2 mRNA signals were widely distributed in the brain, being stronger in the thalamus and in the dopaminergic cells of the mesencephalon. Most brain nuclei displayed both alpha4 and beta2 signals with the exception of some basal ganglia regions and the reticular thalamic nucleus which were devoid of alpha4 signal. alpha6 and beta3 mRNA signals were selectively concentrated in the substantia nigra and the medial habenula. The strongest signals for alpha3 or beta4 mRNAs were found in the epithalamus (medial habenula and pineal gland), whereas there were no specific alpha3 or beta4 signals in mesencephalic dopaminergic nuclei. alpha5 and alpha7 mRNA signals were found in several brain areas, including cerebral cortex, thalamus and substantia nigra, although at a lower level than alpha4 and beta2. The distribution of alpha3, alpha4, alpha5, alpha6, alpha7, beta2, beta3 and beta4 subunit mRNAs in the monkey is substantially similar to that observed in rodent brain. Surprisingly, alpha2 mRNA signal was largely distributed in the Macaca brain, at levels comparable with those of alpha4 and beta2. This observation represents the main difference between rodent and Macaca subunit mRNA distribution and suggests that, besides alpha4beta2*, alpha2beta2* nAChRs constitute a main nAChR isoform in primate brain.     

Postnatal development of the hippocampus in the Rhesus macaque (Macaca mulatta): A longitudinal magnetic resonance imaging study

Nonhuman primates are widely used models to investigate the neural substrates of human behavior, including the development of higher cognitive and affective function. Due to their neuroanatomical and behavioral homologies with humans, the rhesus macaque monkey (Macaca mulatta) provides an excellent animal model in which to characterize the maturation of brain structures from birth through adulthood and into senescence. To evaluate hippocampal development in rhesus macaques, structural magnetic resonance imaging scans were obtained longitudinally at 9 time points between 1 week and 260 weeks (5 years) of age on 24 rhesus macaque monkeys (12 males, 12 females). In our sample, the hippocampus reaches 50% of its adult volume by 13 weeks of age and reaches an adult volume by 52 weeks in both males and females. The hippocampus appears to be slightly larger at 3 years than at 5 years of age. Male rhesus macaques have larger hippocampi than females from 8 weeks onward by approximately 5%. Interestingly, there was increased variability in hemispheric asymmetry for hippocampus volumes at younger ages than at later ages. These data provide a comprehensive evaluation of the longitudinal development of male and female rhesus macaque hippocampus across development from 1 week to 5 years of age. © 2014 Wiley Periodicals, Inc.     

Central nervous system receptors involved in mediating the inhibitory action of neuropeptide Y on luteinizing hormone secretion in the male rhesus monkey (Macaca mulatta)

An earlier finding that gonadotropin-releasing hormone (GnRH) secretion may be triggered prematurely in the juvenile male monkey by central administration of 1229U91, a Y1 receptor antagonist, contributed to our current hypothesis that neuropeptide Y (NPY) is a major component of the brake that holds pulsatile GnRH release in check during prepubertal development in primates. However, 1229U91 is also a Y4 receptor agonist, and the present study was conducted to further examine the role of the Y1 receptor in mediating the putative inhibitory action of NPY on GnRH release. Agonadal juvenile and postpubertal male monkeys were implanted with i.v. and i.c.v. cannulae to gain continuous access to the venous and cerebroventricular circulations without sedation. Luteinizing hormone (LH) secretion was measured to provide an indirect index of GnRH release. The specific Y1 antagonists, VD-11 (476 microg; n = 4) and isopropyl 3-chloro-5-[1-((6-[2-(5-ethyl-4-methyl-1,3-thiazol-2-yl)ethyl]-4-morpholin-4-ylpyridin-2-yl)amino)ethyl]phenylcarbamate (Compound A, 300 microg; n = 4), did not mimic the stimulatory action of 1229U91 on GnRH secretion in the juvenile male monkey. Additionally, neither NPY (200 microg; n = 2), a general Y receptor agonist, nor rPP (100 microg; n = 4), a Y4 agonist, mimicked the action of 1229U91 in stimulating GnRH release. Moreover, previous exposure of the hypothalamus of juvenile monkeys (n = 5) to NPY (660 microg) failed to block 1229U91-induced (200 microg) GnRH release. However, the action of NPY (364 microg) in inhibiting GnRH release postpubertally was attenuated by 1229U91 (300 microg). We conclude that, although the action of exogenous NPY to suppress GnRH release from the postpubertal hypothalamus appears to be mediated, at least in part, by the Y1 receptor, the existence of a Y1 receptor pathway inhibitory to GnRH release in the prepubertal hypothalamus remains to be substantiated.     

Variable Statistical Structure of Neuronal Spike Trains in Monkey Superior Colliculus

Popular models of decision-making propose that noisy sensory evidence accumulates until reaching a bound. Behavioral evidence as well as trial-averaged ramping of neuronal activity in sensorimotor regions of the brain support this idea. However, averaging activity across trials can mask other processes, such as rapid shifts in decision commitment, calling into question the hypothesis that evidence accumulation is encoded by delay period activity of individual neurons. We mined two sets of data from experiments in four monkeys in which we recorded from superior colliculus neurons during two different decision-making tasks and a delayed saccade task. We applied second-order statistical measures and spike train simulations to determine whether spiking statistics were similar or different in the different tasks and monkeys, despite similar trial-averaged activity across tasks and monkeys. During a motion direction discrimination task, single-trial delay period activity behaved statistically consistent with accumulation. During an orientation detection task, the activity behaved superficially like accumulation, but statistically consistent with stepping. Simulations confirmed both findings. Importantly, during a simple saccade task, with similar trial-averaged activity, neither process explained spiking activity, ruling out interpretations based on differences in attention, reward, or motor planning. These results highlight the need for exploring single-trial spiking dynamics to understand cognitive processing and raise the interesting hypothesis that the superior colliculus participates in different aspects of decision-making depending on task differences.SIGNIFICANCE STATEMENT How are decisions based on sensory information transformed into actions? We report that single-trial neuronal activity dynamics in the superior colliculus of monkeys show differences in decision-making tasks depending on task idiosyncrasies and requirements and despite similar trial-averaged ramping activity. These results highlight the importance of exploring single-trial spiking dynamics to understand cognitive processing and raise the interesting hypothesis that the superior colliculus participates in different aspects of decision-making depending on task requirements.     

Effects of early adverse experiences on behavioural lateralisation in rhesus monkeys (Macaca mulatta)

In the past 15 to 20 years, evidence of population-level handedness in non-human primates has emerged from a plethora of studies, although considerable inconsistency is also apparent. The study reported here examined two factors that may contribute to the expression of hand preference: early rearing history and sex differences. Handedness was assessed in rhesus monkeys (Macaca mulatta) using a task that measures coordinated bimanual actions and is referred to as the TUBE task. Nursery-reared monkeys demonstrated greater left-hand bias in the TUBE task when compared to their mother-reared counterparts. Females showed greater right-hand preference and stronger bias on the TUBE task compared to males. These results provide evidence that early rearing experiences significantly influence the development of lateralisation in nonhuman primates.     

Effects of early adverse experiences on behavioural lateralisation in rhesus monkeys (Macaca mulatta)

In the past 15 to 20 years, evidence of population-level handedness in non-human primates has emerged from a plethora of studies, although considerable inconsistency is also apparent. The study reported here examined two factors that may contribute to the expression of hand preference: early rearing history and sex differences. Handedness was assessed in rhesus monkeys (Macaca mulatta) using a task that measures coordinated bimanual actions and is referred to as the TUBE task. Nursery-reared monkeys demonstrated greater left-hand bias in the TUBE task when compared to their mother-reared counterparts. Females showed greater right-hand preference and stronger bias on the TUBE task compared to males. These results provide evidence that early rearing experiences significantly influence the development of lateralisation in nonhuman primates.     

Neurotrophic effects of BDNF on embryonic gonadotropin-releasing hormone (GnRH) neurons

Secretion of gonadotropin-releasing hormone (GnRH) at the median eminence is the essential activator of the reproductive axis. The mechanisms by which embryonic GnRH neurons migrate from the olfactory placode to the preoptic area and then elaborate neurites that course through the hypothalamus to terminate at the median eminence are largely unknown. We investigated the hypothesis that GnRH neurite outgrowth is promoted by brain-derived neurotrophic factor (BDNF) because GnRH neurites course through BDNF-rich areas of the forebrain during their development. Confocal microscopy revealed that most (86%) cultured embryonic GnRH cells tagged with a green fluorescent protein reporter were immunoreactive for TrkB. In primary cultures of E12.5 olfactory tissue, treatment with BDNF induced a dose-dependent increase in neurite outgrowth, but had no discernible effect on branching. BDNF induced phosphorylation of Ca(2+)/cAMP response element-binding protein (pCREB) in both GnRH and non-GnRH cells in these cultures. This was not associated with phosphorylation of ERK in GnRH-immunoreactive cells, though BDNF treatment did stimulate pERK in neighbouring non-GnRH cells. Promotion of neurite outgrowth is unlikely therefore to result from activation of the Ras-MAPK/ERK pathway. We conclude that the developing GnRH secretory system is directly sensitive to BDNF and that this polypeptide functions as a neurotrophic factor for GnRH neurons.     

Early development of forebrain gonadotrophin-releasing hormone (GnRH) neurones and the role of GnRH as an autocrine migration factor

Normal migration of the gonadotrophin-releasing hormone (GnRH) neurones during early development, from the olfactory region to the hypothalamus, is crucial for reproductive development in all vertebrates. The establishment of the GnRH system includes tangential migration of GnRH perikarya as well as extension of GnRH fibres to various areas of the central nervous system (CNS). The exact spatio-temporal nature of this process, as well as the factors governing it, are not fully understood. We studied the development of the GnRH system and the effects of GnRH knockdown using a newly developed GnRH3:EGFP transgenic zebrafish line. We found that enhanced green fluorescent protein is specifically and robustly expressed in GnRH3 neurones and fibres. GnRH3 fibres in zebrafish began to extend as early as 26 h post-fertilisation and by 4–5 days post-fertilisation had developed into an extensive network reaching the optic tract, telencephalon, hypothalamus, midbrain tegmentum and hindbrain. GnRH3 fibres also innervated the retina and projected into the trunk via the spinal cord. GnRH3 perikarya were observed migrating along their own fibres from the olfactory region to the preoptic area (POA) via the terminal nerve ganglion and the ventral telencephalon. GnRH3 cells were also observed in the trigeminal ganglion. The establishment of the GnRH3 fibre network was disrupted by morpholino-modified antisense oligonucleotides directed against GnRH3 causing abnormal fibre development and pathfinding, as well as anomalous GnRH3 perikarya localisation. These findings support the hypothesis that GnRH3 neurones migrate from the olfactory region to the POA and caudal hypothalamus. Novel data regarding the early development of the GnRH3 fibre network in the CNS and beyond are described. Moreover we show, in vivo , that GnRH3 is an important factor regulating GnRH3 fibre pathfinding and neurone localisation in an autocrine fashion.     

Ultrastructural studies of neuronal correlates of the pubertal reaugmentation of hypothalamic gonadotropin-releasing hormone (GnRH) release in the rhesus monkey (Macaca mulatta)

This study tested the hypothesis that puberty in primates is triggered by a remodeling of synaptic inputs and/or glial coverage of hypothalamic gonadotropin releasing hormone (GnRH) neurons. Male rhesus monkeys were prepubertally castrated at 16 months of age and were killed and perfused either 1 month later (n = 4, juvenile group) or at 30 months of age, shortly after initiation of the pubertal increase in pulsatile GnRH release (n = 4, adult group). Hypothalami were sectioned, immunocytochemically stained for GnRH, and processed for electron microscopy. Cross-sectional profiles of 77 GnRH cells from the medial basal hypothalamus (MBH) and the region of the organum vasculosum of the lamina terminalis (OVLT) were compared between the two developmental stages. GnRH cell and nucleolus size in the two groups were the same. The percentage of GnRH perikaryal membrane occupied by synaptic density in the MBH of juveniles was significantly greater (P < 0.05) than that of adults. Differences in the percentage of GnRH perikaryal membrane occupied by synaptic density were not observed in the OVLT nor on GnRH dendrites in either brain region. Qualitative analysis, based on synaptic vesicle shape, failed to reveal developmental differences in putatively excitatory or inhibitory synapses on GnRH cells. The degree of glial ensheathment of GnRH neurons did not change significantly during the two developmental stages. These findings provide ultrastructural evidence for the view that, in primates, neuronal plasticity, and specifically a decrease in synaptic input to GnRH perikarya, may underlie the initiation of the pubertal mode of release of this neuropeptide, and therefore, the onset of puberty in these species. J. Comp. Neurol. 385:71–82, 1997. © 1997 Wiley-Liss, Inc.     

Alzheimer A beta vaccination of rhesus monkeys (Macaca mulatta)

Recent preliminary data suggest that vaccination with Alzheimer A beta might reduce senile plaque load and stabilize cognitive decline in human Alzheimer disease. To examine the mechanisms and consequences of anti-A beta-antibody formation in a species more closely related to humans, rhesus monkeys (Macaca mulatta) were vaccinated with aggregated A beta 1-42. Immunized monkeys developed anti-A beta titers exceeding 1:1000, and their plasma A beta levels were 5- to 10-fold higher than the plasma A beta levels observed in monkeys vaccinated with aggregated amylin. These data support the use of nonhuman primates to model certain phenomena associated with vaccination of humans with aggregated Alzheimer A beta.