Increased galanin synapses onto activated gonadotropin-releasing hormone neuronal cell bodies in normal female mice and in functional preoptic area grafts in hypogonadal mice

Galanin synaptic input onto gonadotropin-releasing hormone (GnRH) neuronal cell bodies was analysed in female mice using the presynaptic vesicle-specific protein, synaptophysin (Syn) as a marker. In the first experiment, forebrain sections from normal ovariectomized ovarian steroid-primed mice exhibiting a surge of luteinizing hormone were processed for immunohistochemical labelling for GnRH, synaptophysin, galanin and Fos. Two representative sections from each brain, one passing through the anterior septum (anterior section) and the other through the organum vasculosum lamina terminalis-preoptic area (posterior section), were analysed under the confocal microscope. None of the GnRH cells analysed in the anterior sections were Fos immunoreactive (IR) or received input from galanin-IR fibres. In contrast, the majority of GnRH cells in the posterior sections analysed were Fos-positive. The number of galanin synapses onto the Fos-positive GnRH cells was significantly higher than that in the Fos-negative cells in this area of the brain, even though the number of Syn-IR appositions was comparable to each other. Transplantation of preoptic area (POA) into the third cerebral ventricle of hypogonadal (HPG) mice corrects deficits in the reproductive system. In the second experiment, synaptic input to GnRH cells was compared between HPG/POA mice with (functional graft) or without (nonfunctional graft) gonadal development. The mean numbers of Syn-IR appositions and galanin synapses per GnRH cell and the proportion of GnRH cells with galanin input were significantly higher in the functional than in the nonfunctional grafts. The results suggest that galanin can act directly on the GnRH cell bodies and may have an important regulatory role on the GnRH system.     

Neurochemical characterization of receptor-expressing cell populations by in vivo agonist-induced internalization: insights from the somatostatin sst2A receptor

Characterization of both neurochemical phenotype of G protein-coupled receptor (GPCR)-expressing cells and receptor compartmentalization is a prerequisite for the elucidation of receptor functions in the central nervous system. However, it is often prevented by the diffuse and homogeneous distribution of receptor immunoreactivity. This is particularly true for the somatostatin (SRIF) sst2A receptor, which is largely distributed in the mammalian brain. By using this receptor as a model, we investigated whether receptor internalization, a biochemical property shared by numerous GPCRs, would reveal sst2A-expressing cell populations in the rat dorsolateral septum (LSD), a region in which SRIF might play an important modulatory role. Thirty minutes to 1 hour after intracerebroventricular injection of the sst2A receptor agonist octreotide, numerous sst2A-immunoreactive neurons and processes became apparent due to intracytoplasmic accumulation of intensely stained granules. Double-immunolabeling experiments with synaptophysin and MAP2 provided evidence that internalized sst2A receptors are predominantly localized in the somatodendritic compartment. Revealing sst2A receptor-expressing cell bodies permitted to analyze their neurotransmitter content. Quantitative analysis demonstrated an extensive overlap (approximately 85%) between SRIF- and sst2A-expressing neuronal populations. Additionally, numerous SRIF-immunoreactive axon-like terminals were found in close apposition with sst2A-positive cell bodies and dendrites. Taken together, these data suggest that the sst2A receptor is predominantly expressed in LSD neurons as a postsynaptic autoreceptor, thus providing novel neuroanatomic clues to elucidate SRIF neurotransmission in this region. More generally, in vivo agonist-induced internalization appears as a rapid and powerful tool for the neurochemical characterization of GPCR-expressing cell populations in the mammalian brain.     

Increased synaptic input to gonadotropin releasing hormone cells in preoptic area grafts that support reproductive development in female hypogonadal mice

Ultrastructural studies have established that gonadotropin releasing hormone (GnRH) neuronal cell bodies receive sparse synaptic input compared to other neuronal cell types. In the present studies, immunocytochemistry for the presynaptic marker synaptophysin, coupled with confocal microscopy, was employed to evaluate whether there was a difference in synaptic input to GnRH cells within preoptic area grafts (hypogonadal, HPG; preoptic area, POA) in hypogonadal female mice that did or did not show ovarian development. GnRH cells in HPG/POA mice with ovarian development exhibited significantly higher numbers of synaptophysin immunoreactive (syn-IR) appositions as compared with HPG/POA mice without ovarian development. This suggests that synaptic input to the grafted GnRH cells is important for the correction of reproductive functions in HPG/POA mice. Following mating, Fos immunoreactivity was present in several GnRH cells in HPG mice with successful POA grafts, indicating the establishment of neuronal projections conveying somatosensory information to the GnRH cells in these mice. The presence of a higher number of syn-IR appositions to GnRH cells in the successful grafts supports this hypothesis.     

Ultrastructural Localization of Synaptophysin to the Secretory Granules of Normal Glucagon and Insulin Cells in Human Islets of Langerhans

Immunogold labeling of the pancreatic islets in humans by means of monoclonal antibodies to synaptophysin resulted in a distinct localization of gold particles to the secretory granules of glucagon-immunoreactive cells. The same type of immunoreactivity was noted with antiserum to chromogranin A. Glucagon immunoreactivity was concentrated in the dense central core of the secretory granules. Some immunoreactivity for synaptophysin was also found in the secretory granules of the insulin-producing cells, although it was weaker in this location.     

Molecular milestones that signal axonal maturation and the commitment of human spinal cord precursor cells to the neuronal or glial phenotype in development.

Insights into the programmatic induction of neuronal and glial genes during human embryogenesis have depended largely on extrapolations of data derived from experimental mammals. However, the assumptions upon which these extrapolations are based have not been rigorously tested. Indeed, practically no information is available even on the human counterparts of the relatively small subset of well-characterized, developmentally regulated neuron and glial specific genes of the mammalian CNS. Thus, the developmental programs upon which human neural embryogenesis are based remain largely undeciphered. We have addressed this problem in immunohistochemical studies conducted on 22 human fetal spinal cords with gestational ages (GAs) that ranged from 6 to 40 weeks by using monoclonal antibodies to several classes of neuron or glial specific polypeptides. These polypeptides included: representatives of four different types (Types I-IV) of intermediate filament proteins, i.e., vimentin filament protein (VFP), glial fibrillary acidic protein (GFAP), different phospho-isoforms of the high (NF-H), middle (NF-M), and low (NF-L) molecular weight (Mr) neurofilament (NF) subunits, both acidic and basic cytokeratin (CK) proteins; three different microtubule associated proteins (MAPs), i.e., MAP2, MAP5, and tau; two different synaptic or coated vesicle proteins, i.e., synaptophysin (SYP) and clathrin light chain B (LCb); an oligodendroglial specific protein, i.e., myelin basic protein (MBP); and a receptor for a CNS trophic factor, i.e., the nerve growth factor receptor (NGFR).(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of Panaxtriol Saponins on synaptophysin and postsynaptic density-95 expression at different periods of cerebral infarction

BACKGROUND:The change in expression of synaptophysin (Syp) and postsynaptic density-95 (PSD-95) alters after cerebral infarction,and the plasticity of synapses contributes greatly to nerve function recovery. Chinese medicinal substances may play an important role in the expression of Syp and PSD-95. OBJECTIVE: To observe the effect of Panaxtriol Saponins (PTS),an active component in Sanqi tongshu capsules,on the expression of Syp and PSD-95 after cerebral infarction at different time points in rats,so as to...     

New Method for Imaging Innervation of the Renal Preglomerular Vasculature. Alterations in Hypertensive Rats

Objective : To develop a new method for viewing adrenergic innervation along renal preglomerular vessels; to assess nerve densities and vascular lesions along arcuate arteries (ArcA), arcuate arterial branches (ArcB), and interlobular arteries (ILA) in spontaneously hypertensive rats (SHR) and in angiotensin II (AngII) and in N^G‐nitro‐l ‐arginine methyl ester (l ‐NAME) hypertensive rats. Methods : Preglomerular vasculatures were isolated after HCl maceration and were immunostained against synaptophysin, a membrane protein of synaptic vesicles. Lesions were stained with Sudan black. Longitudinal nerve densities and relative frequencies of ArcA, ArcB, and ILA endowed with sudanophilic lesions were assessed separately. Results : Synaptophysin immunostaining revealed the vascular neural plexus. Nerves were adrenergic, as the plexus was destroyed by treatment with 6‐hydroxy dopamine. Vascular lesions were not seen in SHR, and increased nerve density was observed along ArcA and ILA. In l ‐NAME‐ and AngII‐hypertensive rats, vascular lesions affected predominantly ArcB and ILA, and nerve density was reduced by 12% and 28% (ArcA), 37% and 31% (ArcB), and by 55% and 34% (ILA), respectively, versus normotensive controls. Endothelin‐1 receptor blockade did not affect AngII‐induced hypertension but prevented both lesion development and reduction of density of the vascular neural plexus. Conclusions : The method we have devised provides a direct en face view of the vascular adrenergic innervation of isolated preglomerular vasculature. Measurements in hypertensive rat models suggest a link between vascular lesions and reduction in nerve density in hypertension. Endothelin‐1 likely plays a key role in mediating both vascular injury and altered vascular nerve density in hypertension.     

Immunolocalization and quantitation of a novel nerve terminal protein in spinal cord development

In the adult spinal cord, the neuron-specific protein NT75 is located in nerve terminals synapsing in the superficial laminae of the dorsal horn. The present study examines the occurrence of NT75 in the developing rat spinal cord. NT75 immunoreactivity is detectable in primary afferent axons at the dorsal root entry zone on embryonic day 15. Subsequently, staining of presumptive nerve terminals appears in the deeper laminae of the dorsal horn, expanding into the superficial laminae during the first postnatal week. NT75 staining also appears in developing corticospinal tract axons in the brainstem at birth, and at lumbosacral levels by postnatal day 5. As NT75-positive nerve terminals approach the adult distribution, staining of primary afferent and corticospinal axons decreases, becoming undetectable by postnatal day 30. Dense transient staining of presumed nerve terminals in the ventral horn is also apparent during early postnatal development. Quantitative analysis of developing spinal cord shows a low level of NT75 immunoreactivity at birth. NT75 activity then increases substantially, reaching values by the third and fourth postnatal weeks up to 2.5 times that seen in adults. The occurrence of NT75 immunoreactivity correlates with the reported time course of synaptic development in the spinal cord. In addition, the results suggest that NT75 immunoreactivity is maintained at high levels in the nerve terminals of certain neural pathways into adulthood, whereas in other systems NT75 immunoreactivity may be detectable only during development.     

转基因小鼠脑组织中突触素和发动蛋白Ⅰ及衔接蛋白180表达的变化

目的探讨瑞典型淀粉样前体蛋白突变基因转基因小鼠脑组织中突触素(synaptophysin)、发动蛋白Ⅰ(dynamin Ⅰ)及衔接蛋白180(AP180)表达变化。方法选择6只瑞典型淀粉样前体蛋白突变基因转基因小鼠为转基因组,另选5只小鼠为对照组。采用免疫组织化学染色法检测小鼠海马及颞叶皮质synaptophysin、dynamin Ⅰ及AP180的表达,图像分析半定量;免疫组织化学双染法观察转基因小鼠脑组织中synaptophysin与β淀粉样蛋白(Aβ)_(1-42)在老年斑表达部位的关系。结果与对照组比较,转基因组小鼠脑组织齿状回分子层、海马CA1、CA3及内嗅区皮质各层synaptophysin平均灰度值明显增高(P0.05,P0.01);齿状回颗粒细胞、海马CA1锥体细胞及内嗅区皮质各层dynamin Ⅰ平均灰度值明显增高(P0.05,P0.01);齿状回分子层、海马CA4、CA1、内嗅区皮质各层及颞叶皮质Ⅱ～Ⅴ层AP180平均灰度值明显增高(P0.05,P0.01)。免疫组织化学双染显示转基因组小鼠老年斑内有synaptophysin和Aβ_(1-42)共同存在。结论转基因小鼠脑组织中synaptophysin、dynamin Ⅰ及AP180表达降低,提示出现不同程度突触丧失及突触囊泡回收功能缺陷,可能是该小鼠认知功能障碍的原因之一。