A peptide derived from a neurite outgrowth-promoting domain on the gamma 1 chain of laminin modulates the electrical properties of neocortical neurons

Laminins form a family of large multidomain glycoproteins of the extracellular matrix. The cellular distribution of laminin immunoreactivity in the adult mammalian central nervous system suggests an important role for laminins in mature brain function in addition to their role during brain development. To characterize the effects of this group of extracellular matrix molecules on mature brain function, intracellular recording techniques were applied to in vitro slice preparations of the rat neocortex. The experiments show that a peptide homologous to the C-terminal part of the gamma 1 chain of laminin modulates the electrical activity of pyramidal neurons in the adult neocortex of the rat. The peptide is part of the neurite outgrowth-promoting domain of the gamma 1 chain on the E8 fragment of laminin and it displays the neurite outgrowth-promoting activity of the native laminin molecule. Perfusion of in vitro brain slices with the peptide increased the input resistance of the neuronal membrane. In addition, a rise in inward rectification could be observed. These events were accompanied by a strong increase in direct excitability of the treated neurons. Immunohistochemistry techniques were applied to sections of the adult rat neocortex and hippocampus to demonstrate the presence of both the neurite outgrowth-promoting domain and the native laminin in the adult brain. An antiserum raised against the neurite outgrowth-promoting domain on the gamma 1 chain of laminin, which also recognized the free synthetic peptide, showed immunoreactivity on neurons. In addition, a population of glial fibrillary acidic protein-positive astrocytes in the hippocampus displayed immunoreactivity for this antibody. These results were confirmed by using several antibodies directed against the whole laminin-1 molecule. Neurons in the neocortex and hippocampus, as well as astrocytes in the hippocampus, demonstrated immunoreactivity for antibodies directed against the whole laminin-1 molecule. The results suggest that laminins containing the gamma 1 chain have the potential to modulate neuronal activity. This effect may be mediated either by direct cell-cell contact from surrounding cells, or through the neuronal expression of laminin or laminin-like molecules which are inserted into the neuronal cell membrane.     

Id1, Id2, and Id3 gene expression in neural cells during development

Id1, Id2, and Id3 mRNA are expressed mainly in the proliferating ependymal cell zone of the mouse brain during embryogenesis. In this study, the expression pattern and cell phenotypes of the Id family mRNA were examined in postnatal and adult rat brain. The expression of Idl and Id3 mRNA in rat brain was observed in the cortex layer 1, corpus callosum, ventricular/subventricular zone (VZ/ SVZ), and the CA1-4 layers of the hippocampus at postnatal day 1 (P1) through P14, whereby it declined at 2 months. In general, the developmental pattern of Idl mRNA coincided with the pattern observed for Id3 mRNA. Similar to Id1 and Id3, Id2 mRNA was highly expressed in the corpus callosum, VZ/SVZ, and the hippocampus. Examination of Id2 mRNA revealed high levels in the cortex and caudate putamen at P1 through P14, whereas a decline was observed in its expression in the adult cortex. In P5 rat cerebellum, all Id mRNA examined were found in the internal granular cell layers; however, at this time point, only Id2 mRNA expression was detected in the differentiating zone of the external granular cell layers, preferentially localizing to adult Purkinje cells. Furthermore, only Id2 mRNA expression in brain was observed in NF+ neurons at P5. Examination of S100alpha+ and GFAP+ astrocytes, revealed the presence of all three mRNAs, whereas the expression of Id2 and Id3 mRNA was absent in 04+ immature oligodendrocytes. These data suggest that the spatial and temporal kinetic patterns during development, as well as cellular specificity, of the Id gene family may play a critical role in neural precursor cell proliferation and cell divergence.     

Presence of physiologically stimulated RET in adult rat brain: induction of RET expression during nerve regeneration

The product of the RET proto-oncogene is a protein belonging to the receptor-like tyrosine kinase superfamily. RET is expressed in several neural crest-derived cell lineages and has been implicated in the correct development of the peripheral nervous system. To gain further insight into RET function, we investigated the presence of active RET in adult rat tissues. We show, by immunoblotting, that the products of the RET proto-oncogene (p155ret) are present in specific regions of adult rat brain, including the cerebellum, striatum, brainstem, hypothalamus, hippocampus, and olfactory bulb. Moreover, in the cerebellum, p155ret is phosphorylated in tyrosine residues, thus indicating that this brain structure contains p155ret in an activated state. Finally, the presence of RET in motoneurons prompted us to analyze the effects of hypoglossal nerve section on its expression. We observed a dramatic increase in p155ret in the motoneuron nuclei, thus suggesting that RET tyrosine kinase plays a role in the neuronal response to axotomy and/or during nerve regeneration.     

Differential induction of nerve growth factor and basic fibroblast growth factor mRNA in neonatal and aged rat brain

Stimulation of glucocorticoid or beta-adrenergic receptors (BAR) has been shown to increase nerve growth factor (NGF) biosynthesis in adult rat brain. Little is known about the role of these receptors in the regulation of NGF expression in neonatal and aged brain. We have examined the effect of the synthetic glucocorticoid dexamethasone (DEX) and the BAR agonist clenbuterol (CLE) on the levels of NGF mRNA in neonatal (8 day old), adult (3 month old) and aged (24 month old) rats. By 3 h, DEX (0.5 mg/kg, s.c.) evoked a comparable increase in NGF mRNA in the cerebral cortex and hippocampus in both 8-day and 3-month-old rats. In contrast, CLE (10 mg/kg, i.p.) failed to change NGF mRNA levels in neonatal rats, while increasing (2-3-fold) NGF mRNA levels in the cerebral cortex of adult rats. In 24-month-old rats, both DEX and CLE elicited only a modest increase in NGF mRNA. This increase was, however, anatomically and temporally similar to that observed in adult animals. The weak effect of DEX or CLE was not related to a down-regulation of receptor function because both DEX and CLE were able to elicit a comparable increase in the mRNA levels for basic fibroblast growth factor (FGF2) in neonatal, adult and aged rat brain. Our data demonstrate that induction of NGF expression by neurotransmitter/hormone receptor activation varies throughout life and suggest that pharmacological agents might be useful tools to enhance trophic support in aging.     

Neurotrophins and their receptors in the adult hypo- and hyperthyroid rat after kainic acid injection: an in situ hybridization study

Thyroid hormone plays a key role in trophic events during development of the central nervous system. In spite of neurological and psychiatric symptoms associated with adult hypothyroidism, the role of thyroid hormone in mature brain function is less clear. In this paper we investigated the effect of thyroid status on kainic acid-induced up-regulation of mRNAs for members of the nerve growth factor family and related receptors in adult male rats by means of in situ hybridization. We found that in hypothyroid rats there is a dramatic attenuation of the kainic acid-induced up-regulation of mRNA levels for nerve growth factor, brain-derived neurotrophic factor and tyrosine kinase trkB in euthyroid rats. A trend to reduced c-fos mRNA up-regulation, which did not reach significance, was also found, whereas the increase in c-jun mRNA after kainic acid was similar in eu-, hypo- and hyperthyroid rats. These data indicate a severe impairment of the regulation of neurotrophin synthesis after excitotoxin administration in the hippocampus during adult hypothyroidism. Possible roles of thyroid hormone in molecular, biochemical and metabolic mechanisms of this defect are discussed.     

Autoantigen Ku in the brain. Developmentally regulated expression and subcellular localization

A double-stranded DNA end-binding factor with high levels of expression in brain and testis of adult mice was identified as the Ku protein, earlier described as an autoantigen in connective tissue diseases and found to be essential for recombination of the immunoglobulin genes and DNA repair. High Ku levels were found in the cerebellum and pituitary gland, lower levels in the hippocampus, hypothalamus and white matter structures. Ku levels were much higher in embryonic rat brain than in the adult brain, suggesting a role of the Ku protein in brain development. In embryonic rat brain, Ku was associated with cell nuclei, but was predominantly located in the cytosol in the adult rat cerebellum and hippocampus. The abundant expression of Ku in the brain suggests the involvement of Ku autoantibodies in the pathogenesis of neuropsychiatric complications in connective tissue diseases.     

Subcellular distribution of GLUT4 in Chinese hamster ovary cells overexpressing mutant dynamin: evidence that dynamin is a regulatory GTPase in GLUT4 endocytosis

Previous studies have suggested that the neurotrophins brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) are neuroprotective or neurotrophic for certain subpopulations of hippocampal neurons following various brain insults. In the present study, the expression of BDNF and NT-3 mRNAs in rat hippocampus was examined after traumatic brain injury. Following lateral fluid percussion (FP) brain injury of moderate severity (2.0-2.1 atm) or sham injury, the hippocampi from adult rats were processed for the in situ hybridization localization of BDNF and NT-3 mRNAs using 35S-labeled cRNA probes at post-injury survival times of 1, 3, 6, 24 and 72 h. Unilateral FP injury markedly increased hybridization for BDNF mRNA in the dentate gyrus bilaterally which peaked at 3 h and remained above control levels for up to 72 h post-injury. A moderate increase in BDNF mRNA expression was also observed bilaterally in the CA3 region of the hippocampus at 1, 3, and 6 h after FP injury, but expression declined to control levels by 24 h. Conversely, NT-3 mRNA was significantly decreased in the dentate gyrus following FP injury at the 6 and 24 h survival times. These results demonstrate that FP brain injury differentially modulates expression of BDNF and NT-3 mRNAs in the hippocampus, and suggest that neurotrophin plasticity is a functional response of hippocampal neurons to brain trauma.     

Ontogenic expression of natriuretic peptide mRNAs in postnatal rat brain: implications for development?

The central natriuretic peptide system is composed of at least three structurally homologous and uniquely distributed peptides and receptors which are thought to be involved in the central regulation of cardiovascular and autonomic function and more recently been shown to affect cellular growth and proliferation, processes pertinent to mammalian development. As such, following our initial mapping of preproatrial natriuretic peptide (ppANP) mRNA in adult brain [M.C. Ryan, A.L. Gundlach, Anatomical localization of preproatrial natriuretic peptide mRNA in the rat brain by in situ hybridization histochemistry: in olfactory regions, J. Comp. Neurol., 356 (1995) 168-182], it was of interest to determine the ontogenic expression of natriuretic peptide mRNAs in the developing rat brain. Using in situ hybridization histochemistry of specific [35S]- or [33P]-labeled oligonucleotides, ppANP and preproC-type natriuretic peptide (ppCNP) mRNAs were detected in the developing rat brain from postnatal day 4 to day 60 (adult). PpANP mRNA was observed in many hindbrain, but only some forebrain, regions at postnatal day 4. Regional differences in the temporal expression of ppANP mRNA were apparent with ppANP mRNA detected in the medial preoptic area, mammillary nuclei and medial habenular nucleus at postnatal day 4 and in other areas including the arcuate and dorsomedial hypothalamic nuclei and in olfactory and limbic regions at postnatal day 10. A number of regions also exhibited transient expression of ppANP mRNA such as the bed nucleus of the stria terminalis and the medial cerebellar nucleus. In contrast, ppCNP mRNA was detected at relatively high levels in several regions on postnatal day 4 including olfactory nuclei, the hippocampus and particularly the pontine nucleus. The level of expression appeared to increase markedly in most regions including forebrain olfactory and hippocampal areas and in brainstem regions including the pontine nucleus, the parvocellular and lateral reticular and spinal trigeminal nuclei by postnatal days 10 and 13, but decreased from this peak to equivalent to adult levels by postnatal day 28. The differential and transient expression of the natriuretic peptides during postnatal development, together with previous reports of the ontogenic regulation of natriuretic peptide receptor expression and binding patterns, further suggests their involvement in developmental processes in the rat CNS and provides information relevant to the likely functional development of natriuretic peptide-utilizing pathways.     

Postnatal expression of H1-receptor mRNA in the rat brain: correlation to L-histidine decarboxylase expression and local upregulation in limbic seizures

Histamine is implicated in the regulation of brain functions through three distinct receptors. Endogenous histamine in the brain is derived from mast cells and neurons, but the importance of these two pools during early postnatal development is still unknown. The expression of histamine H1-receptor in the rat brain was examined using in situ hybridization during postnatal development and in adults. For comparison, the expression of L-histidine decarboxylase (HDC) in the two pools was revealed. H1-receptor was evenly expressed throughout the brain on the first postnatal days, but resembled the adult, uneven pattern already on postnatal day 5 (P5). HDC was expressed in both mast cells and tuberomammillary neurons from birth until P5, after which the mast cell expression was no more detectable. In adult rat brain, high or moderate levels of H1-receptor expression were found in the hippocampus, zona incerta, medial amygdaloid nucleus and reticular thalamic nucleus. In most areas of the adult brain the expression of H1-receptor mRNA correlates well with binding data and histaminergic innervation. A notable exception is the hypothalamus, with high fibre density but moderate or low H1-receptor expression. Systemic kainic acid administration induced increased expression of H1-receptor mRNA in the caudate-putamen and dentate gyrus, whereas no change was seen in the hippocampal subfields CA1-CA3 or in the entorhinal cortex 6 h after kainic acid injections. This significant increase supports the concept that histaminergic transmission, through H1-receptor, is involved in the regulation of seizure activity in the brain.     

The cDNA cloning and ontogeny of mouse alpha-synuclein

Alpha-synuclein has been implicated in the pathogenesis of Parkinson's disease. To investigate the role of alpha-synuclein in the brain, the cDNA clone encoding the mouse cognate of the human alpha-synuclein was isolated from a mouse brain cDNA library. The open reading frame coded for 140 amino acids that share 95% identity with human alpha-synuclein. Northern blot analysis showed that alpha-synuclein mRNA was primarily expressed in brain and spleen of adult mouse. In situ hybridization histochemistry revealed the highest expression of alpha-synuclein mRNA in the hippocampal formation and neocortex of the adult mouse. alpha-Synuclein mRNA expression in the brain was first observed in the hippocampus and neocortex on postnatal day 1. Levels of alpha-synuclein mRNA in these forebrain areas were nearly maximal at postnatal day 7 and remained relatively high until the adult stage. alpha-Synuclein mRNA was expressed in the liver transiently during embryogenesis.     

Influence of renal failure on the pharmacokinetics and neuromuscular effects of a single dose of rapacuronium bromide

Neurotransmitter transporters are involved in termination of the synaptic neurotransmission and are implicated as the sites of action of antidepressant medicines and illicit drugs. In addition to their function in neurotransmission, neurotransmitter transporters play a key role in neuroregulation and brain development. In this report, the developmental distribution of the "orphan" transporter NTT4, whose substrate has not yet been shown, is described. Immunohistochemical studies have previously shown NTT4 to be specifically and widely localized to the central nervous system. In this report, the distribution of NTT4 in brain areas enriched in glutamatergic and gamma-aminobutyric acid-ergic innervations is further substantiated. NTT4 is detected beginning at E18 in various parts of the rat brain, including the cerebral cortex, fimbria hippocampi, fornix, lateral lemniscus, anterior commissure, and spinal cord. At E18, strong immunoreactivity of NTT4 is observed in the cortical subplate and marginal layers that later develops into the fimbria hippocampi, and at P22, the expression of NTT4 in the hippocampal formation reaches the mature form. The expression of NTT4 in the spinal cord begins at E18 in the ventral white matter. Heavy staining for NTT4 is observed in the substantia nigra since birth and through all time points examined. Transient immunoreactivity is observed in the inferior colliculus, reaching maximal expression at P10, whereas the superior colliculus commences to express NTT4 only after this time point. The globus pallidus is highly stained after birth, and the caudate putamen shows strong staining for NTT4 only at P22. In the adult rat brain, NTT4 is strongly expressed in the olfactory bulb, cerebral cortex, striatum, hippocampus, thalamus, substantia nigra, pontine nucleus, cerebellum, and spinal cord. The developmental distribution of NTT4 suggests involvement in central nervous system maturation.