PubMed Health⌕ Search

Biomedical subjects

L I Benowitz

Publications and source records attributed to L I Benowitz.

At least 55 records · Page 3Linked to original sources
PubMed1990

Immunoreactive GAP-43 in the neuropil of adult rat neostriatum: localization in unmyelinated fibers, axon terminals, and dendritic spines.

GAP-43 is a neuron-specific phosphoprotein that has been implicated in neuronal development, axonal regeneration, and synaptic plasticity. Although in mammals the caudate-putamen is among those brain areas that retain a high content of GAP-43 throughout life, the role of the phosphoprotein in the neostriatum is unknown. In order to understand better the possible function(s) of GAP-43 in the adult striatum, its cellular localization was examined with immunohistochemistry at the light and electron microscopic levels by using a sheep polyclonal antibody. At the light microscopic level immunoreactive GAP-43 was abundant throughout the neostriatal neuropil but was absent from neuronal somata. At the ultrastructural level, labeling was most prevalent in small unmyelinated axons (0.12-0.15 microns diameter). Reaction product was distributed along fibers in discrete patches about 1 micron apart and in preterminal sites from which vesicle-filled boutons arose. Staining was also present in small (0.35 microns) axon terminals that contained round vesicles and formed asymmetric synapses, mostly with thin spines. Following unilateral cortical lesions, some degenerating cortical axons in the neostriatum exhibited GAP-43 labeling. Unexpectedly, in normal striatum, GAP-43 was also occasionally found in the heads of dendritic protrusions and in thin spines that received asymmetric contacts. We speculate that in the adult neostriatum, the protein may be important in the remodeling of synapses onto medium spiny neurons that involve, in part, the corticostriatal pathway.

Animals↗
PubMed1990

Abnormal retinal projections alter GAP-43 patterns in the diencephalon.

In Syrian hamsters, mature retinal terminals contain only low levels of the growth-associated protein, GAP-43, whereas the lateral posterior nucleus (LP) of the thalamus contains high levels of this protein. Damage to the superior colliculus in neonatal hamsters induces retinal terminals to form dense patches of innervation in the LP, an area which otherwise receives little if any direct retinal input. The present study used GAP-43 antibodies to examine the interaction between abnormally routed optic fibers and the cells in the anomalous thalamic target zone. Immunohistochemistry revealed very little GAP-43 in the abnormal retinal projection to the LP, indicating that the normal developmental decline in GAP-43 levels occurs even in an inappropriate extracellular environment. Moreover, retinal fibers were found to exclude the protein from its normal territory, forming negatively-stained islands in those regions of the LP containing the retinal terminals. In order to identify the normal source of GAP-43-positive terminals in the LP, we surgically removed two major extrinsic afferents to this region, or we chemically eliminated local interneurons. Whereas removing projections from the SC or posterior cortex did not alter GAP-43 immunoreactivity in the LP, destruction of local interneurons with ibotenic acid resulted in markedly diminished levels of this protein. These results show that retinal terminals induced to form in an abnormal target area undergo their normal diminution of GAP-43, and that these retinal projections displace other GAP-43-rich terminals in the LP that appear to arise from local interneurons.

Animals↗
PubMed1990

Transient patterns of GAP-43 expression during the formation of barrels in the rat somatosensory cortex.

The development of the rat barrel field cortex was investigated with an antibody to the axonal membrane-specific phosphoprotein GAP-43 in order to examine the developmental pattern of afferent projections, and with cytochrome oxidase histochemistry and Nissl stains to reveal the morphogenesis of cortical barrels. On the first two days after birth, GAP-43 immunostaining in the cortical plate was light and diffuse, then became intense in the presumptive layer IV of the parietal cortex on PND3 (day of birth = PND0). Immunoreactive densities were visible as small, focal patches within the centers of prospective barrels. These densities increased in size and intensity over the next few days and then diminished abruptly. On PND7, the distribution of GAP-43 was coextensive with barrels, as defined by cytochrome oxidase histochemistry and Nissl staining. GAP-43 virtually disappeared from the barrels after PND7. From the second postnatal week, GAP-43 immunostaining was evident in the septa between barrels and in the dysgranular regions of SI cortex. This pattern of GAP-43 distribution was complementary to the pattern of cytochrome oxidase activity, and persisted into maturity. In an attempt to identify possible source(s) of GAP-43 positive afferents in the developing barrels, we examined the effects of altering the sensory periphery on the distribution of GAP-43 immunostaining in the cortex. Rat pups had row C whiskers cauterized on PND0 and were sacrificed on PND3 or PND5. Whereas immunopositive densities corresponding to intact whiskers developed in a normal, punctate pattern, cortical representation of the lesioned whiskers formed a continuous band of labeling that was evident as early as PND3. We argue that the disjunctive expression of GAP-43 in the barrel field reflects the pattern of distribution of afferents (most likely from the ventro-basal thalamic nucleus) to the barrel field cortex, and that this pattern may be instructive in the formation of barrels as cytoarchitectonic units. The rapid alteration in patterns of immunostaining following whisker lesions lends further support to the conclusion that the "barrel template" is conveyed to the neocortex by incoming afferents. The possible significance of the transient expression of GAP-43 in the maturing barrel field is discussed.

Aging↗
PubMed1990

Impaired verbal reasoning and constructional apraxia in subjects with right hemisphere damage.

In addition to causing visuospatial deficits, damage to the right cerebral hemisphere also impairs other cognitive abilities, including those requiring higher-order aspects of language. The present study used a standardized test battery to examine the relationship between visuospatial abilities and comprehension of narrative material in subjects having unilateral right hemisphere damage (RHD). In a series of 41 consecutively admitted RHD subjects, impairments in abstracting information from narrative passages were as prevalent and as severe in magnitude as constructional apraxia. Moreover, the extent of the visuospatial and linguistic impairments were highly correlated. Although age, educational levels, and degree of premorbid brain atrophy were all found to influence performance, analysis of a select subgroup of the population established that the covariation of visuospatial and verbal impairments is related to right hemisphere damage per se. Clinically, these findings may be of significance for understanding the pervasive cognitive impairments that are often evidenced by RHD patients.

Aged↗
PubMed1990

The pattern of GAP-43 immunostaining changes in the rat hippocampal formation during reactive synaptogenesis.

The reactive synaptogenesis that takes place in the rat hippocampal formation after certain experimental manipulations affords an opportunity to investigate the molecular events that underlie structural remodeling in the adult CNS. Between 2 and 4 days after lesioning the perforant pathway, levels of the synaptic phosphoprotein, GAP-43 (B50, F1, pp46, neuromodulin), were found to increase markedly in the inner molecular layer (iml) of the dentate gyrus, coincident with the time at which commissural-associational (CA) fibers begin to sprout axon collaterals into dendritic portions denervated by the lesion. GAP-43 immunostaining in the iml began to decline by 8 days but continued to define an expanded CA projection for at least one month. In the outer molecular layer (oml), GAP-43 levels decreased after the loss of perforant pathway terminals and did not return for 2-3 weeks, the time at which sprouting of septal inputs into this layer can be visualized by cholinesterase histochemistry. These results demonstrate that GAP-43 levels change during reactive synaptogenesis, and point to differences among neural systems in their expression of this protein.

Animals↗
PubMed1990

Transfection of PC12 cells with the human GAP-43 gene: effects on neurite outgrowth and regeneration.

The neuronal growth associated protein GAP-43 is expressed at high levels during axonal growth and regeneration. In this report, we describe the transfection of the nerve growth factor (NGF)-responsive pheochromocytoma cell line PC12 with the human GAP-43 cDNA under the control of the Moloney murine leukemia virus long terminal repeat (MoMuLV LTR). Two PC12 subclones were isolated that constitutively expressed GAP-43 from the transfected cDNA and showed increased responsiveness to NGF. Of the two transfected PC12 subclones, the subclone expressing the most human GAP-43 RNA showed an accelerated initial neurite outgrowth response and a 10-fold increased sensitivity to NGF. Neurite regeneration was significantly enhanced in both transfected subclones and, in contrast to untreated PC12 cells, could occur transiently in the absence of added NGF. These results suggest that GAP-43 may potentiate the action of NGF on neurite initiation and regeneration.

Cell Differentiation↗
PubMed1990

The growth-associated protein GAP-43 appears in dorsal root ganglion cells and in the dorsal horn of the rat spinal cord following peripheral nerve injury.

When adult dorsal root ganglion cells are dissociated and maintained in vitro, both the small dark and the large light neurons show increases in the growth-associated protein GAP-43, a membrane phosphoprotein associated with neuronal development and plasticity. Immunoreactivity for GAP-43 appears in the cytoplasm of the cell bodies as early as 3.5 h post axotomy and is present in neurites and growth cones as soon as they develop. At early stages of culture (4 h to eight days) satellite/Schwann cells are also immunoreactive for GAP-43. Neurons in isolated whole dorsal root ganglion maintained in vitro become GAP-43-immunoreactive between 2 and 3 h after axotomy. It takes three days however, after cutting or crushing the sciatic nerve in adult rats in vivo, for GAP-43 immunoreactivity to appear in the axotomized dorsal root ganglion cells. GAP-43 immunoreactivity can be detected in the central terminals of primary afferent neurons in the superficial laminae of the dorsal horn of the lumbar enlargement four days after sciatic cut or crush. The intensity of the GAP-43 staining reaches a peak at 21 days and becomes undetectable nine weeks following crush injury and 36 weeks following sciatic nerve cut. The pattern of GAP-43 staining is identical to the distribution of sciatic small-calibre afferent terminals. Little or no staining is present in the deep dorsal horn, but GAP-43 does appear in the ipsilateral gracile nucleus 22 days after sciatic injury. In investigating the mechanism of GAP-43 regulation, blockade of axon transport in the sciatic nerve with vinblastine (10(-5) M-10(-4) M) or capsaicin (1.5%) was found to produce a pattern of GAP-43 immunoreactivity in the dorsal horn identical to that found with crush, while electrical stimulation of the sciatic nerve had no effect. Axotomy of primary sensory neurons or the interruption of axon transport in the periphery therefore acts to trigger GAP-43 production in the cell body. The GAP-43 is transported to both the peripheral and the central terminals of the afferents. In the CNS the elevated GAP-43 levels may contribute to an inappropriate synaptic reorganization of afferent terminals that could play a role in the sensory disorders that follow nerve injury.

Animals↗
PubMed1989

Immunohistochemical localization of GAP-43 in the developing hamster retinofugal pathway.

Metabolic labeling studies have shown that the developing hamster retinotectal pathway is marked by a high level of synthesis and axonal transport of the neuron-specific phosphoprotein GAP-43, which then decline sharply with synaptic maturation. To understand better the relationship of GAP-43 to specific developmental events, we used a monospecific antibody to examine the location of this protein in the optic tract and retinal target areas at various stages. In late embryonic and in neonatal hamsters, dense GAP-43 immunostaining was seen along the entire extent of the optic tract axons, including fascicles coursing over and through the lateral geniculate body (LGB) and within the upper layers of the superior colliculus (SC). The retinal origin of many of these fascicles was confirmed by their rapid disappearance after removal of the contralateral eye. During the first postnatal week, immunostaining in the fiber fascicles showed a marked decline, though the protein was still present throughout the neuropil of the LGB and SC. In the second postnatal week, the neuropil staining also diminished, and by 12 days after birth, both structures showed only light immunoreactivity. The high levels of GAP-43 in embryonic and neonatal optic tract axons coincide temporally with axon elongation, initial target contact, and collateral formation by the retinofugal fibers, whereas subsequent concentration of the protein in the neuropil suggests its involvement in the elaboration of terminal arbors and synaptogenesis.

Aging↗
PubMed1989

Peripheral nerve regeneration induces elevated expression of GAP-43 in the brainstem trigeminal complex of adult hamsters.

A polyclonal antibody was used to delineate the pattern of GAP-43 expression in central processes of trigeminal ganglion cells while their peripheral processes were undergoing regeneration. Two weeks after transection of the infraorbital nerve, levels of GAP-43 ipsilateral to the transection were greatly increased along the trajectory of infraorbital axons in the central trigeminal tract and also within the target neuropil. We conclude that elevated levels of GAP-43 in central processes of injured trigeminal ganglion cells occur in direct response to the regenerative response of the cell body and may have important implications for 'plasticity'-related changes seen in the adult trigeminal system.

Animals↗
PubMed1989

The amyloid precursor protein is concentrated in neuronal lysosomes in normal and Alzheimer disease subjects.

The 4.2-kilodalton (kDa) polypeptide associated with the cerebral amyloid deposits of Alzheimer's disease (AD) derives from a much larger protein that is encoded by a gene on chromosome 21. In the present study, we have used antibodies raised against portions of the amyloid protein precursor (APP) to map its normal distribution and to gain further insights into the events that lead to amyloid deposition. Antibodies raised against several different portions of APP reacted with proteins having apparent molecular sizes of 65, 67, and 132 kDa on Western blots. In sections through the normal human brain, immunocytochemistry revealed punctate concentrations of the protein in pyramidal cells of the neocortex, particularly in associative regions, and intense staining in the CA1 pyramidal cells of the hippocampus. By electron microscopy, this punctate distribution coincided with dense concentrations of the protein in secondary lysosomes. In the hippocampus of several AD cases examined, abnormally dense immunostaining in enlarged intracellular domains accompanied a severe atrophy of the CA1 neurons. These data suggest that accumulations of APP in lysosomes of particular neurons may, in AD, lead to proteolytic events that form the insoluble 4.2-kDa amyloid peptide.

Aged↗
PubMed1989

Partial purification and characterization of a neurite-promoting factor from the injured goldfish optic nerve.

We have partially purified and characterized a neurite-promoting factor derived from the injured goldfish optic nerve (ON). This factor is secreted into conditioned media (CM) by the injured, but not intact goldfish ON, and has potent outgrowth-promoting effects on neurons of the embryonic mammalian brain. Based on its elution properties on ion-exchange and gel-filtration chromatography, this factor appears to be an acidic protein of Mr ca. 26 kilodaltons (kDa) that is distinct from previously characterized growth factors with described effects on mammalian CNS neurons.

Animals↗
PubMed1989

Protein fatty acid acylation in developing cortical neurons.

Neuron-enriched cultures derived from embryonic day 17 rat cerebral cortex were incubated in the presence of [3H]myristic or [3H]palmitic acid. Analysis of radiolabeled proteins by two-dimensional gel electrophoresis and fluorography revealed extensive incorporation of fatty acids into a small number of neuronal proteins. The major acylated proteins had apparent molecular weights and isoelectric points as follows: 87,000, 4.3; 63,000, 4.4; 45,000, 4.4; and 20,000, 5.3. After labeling with [3H]myristic acid, the radioactivity associated with these proteins was identified as myristic acid, which was attached via an ester linkage. All four of the major acylated neuronal proteins were found to be membrane-bound and enriched in growth cones. By virtue of its molecular weight, isoelectric point, subcellular distribution, and peptide map, the 87-kilodalton polypeptide was shown to be equivalent to pp80, a phosphoprotein that has been described in developing and mature synaptic terminals. The 45-kilodalton acylated protein also appears to coincide with another growth cone phosphoprotein, pp40. Acylation may serve as a mechanism to regulate the function of these proteins, or may play a role in directing them to the nerve terminal membrane.

Acylation↗
PubMed1989

Localization of the growth-associated phosphoprotein GAP-43 (B-50, F1) in the human cerebral cortex.

The growth-associated phosphoprotein GAP-43 is a component of the presynaptic membrane that has been linked to the development and functional modulation of neuronal connections. A monospecific antibody raised against rat GAP-43 was used here to study the distribution of the protein in cortical and subcortical areas of the human brain. On Western blots, the antibody recognized a synaptosomal plasma membrane protein that had an apparent molecular weight and isoelectric point similar to GAP-43 of other species. In brain tissue reacted with the antibody, the heaviest immunoreactivity was found in associative areas of the neocortex, particularly within layers 1 and 6, in the molecular layer of the dentate gyrus, the caudate putamen, and the amygdala. In contrast, primary sensory or motor regions of the cortex, portions of dorsal thalamus, and cerebellum showed only light staining. Staining was generally confined to the neuropil, which showed punctate labeling, whereas most neuronal somata and fiber bundles were unreactive. The pronounced variations in GAP-43 immunostaining among various areas of the human brain may reflect different potentials for functional and/or structural remodeling.

Cerebral Cortex↗
PubMed1988

A factor from the injured lower vertebrate CNS promotes outgrowth from human fetal brain neurons.

Unlike mammals, lower vertebrates retain the capacity to regenerate damaged central nervous system (CNS) pathways throughout life. In previous studies, we have used the goldfish optic nerve (ON) as a model for CNS regeneration, and found that the injured goldfish ON selectively secretes a factor that promotes process outgrowth of cultured neurons, including neurons of the developing rodent CNS. In the current study, we found that a factor similarly obtained from the injured goldfish ON also has potent outgrowth-promoting effects on cerebrocortical neurons of the fetal human brain, and that these effects are dependent on the age of fetal neurons. This factor appeared to be a protein of mol. wt. greater than 12,000, and was associated with a distinctive morphology of neurite outgrowth. The neurite-promoting factor from the injured goldfish ON may be homologous to factors within the developing human brain.

Animals↗
PubMed1988

Extraction of major acidic Ca2+ dependent phosphoproteins from synaptic membranes.

The association of several phosphoproteins with the synaptosomal plasma membrane (SPM) was investigated by phosphorylating SPM fractions from neonatal rat brain in the presence of Ca2+ and then exposing these to a variety of agents. Extraction of the major acidic phosphoproteins, GAP-43, pp40, and pp80, was assessed by two-dimensional gel electrophoresis and fluorography. All three proteins were best extracted from the membrane by high pH and by guanidine hydrochloride. GAP-43 was not extracted in the presence of either low- or high-ionic-strength buffers, reducing agents, or chelating agents; pp80 and pp40, however, showed a significant extraction even under low-ionic-strength conditions. Partition experiments with Triton X-114 revealed an amphiphilic behavior for GAP-43 and a strong affinity for hydrophobic environments for pp80 and pp40. None of the phosphoproteins was released from the membrane by the use of a phosphatidylinositol-specific phospholipase C. The extraction properties of GAP-43, pp80, and pp40 are similar to those of known extrinsic membrane proteins and therefore suggest that these phosphoproteins are peripheral rather than integral to the membrane compartment.

Animals↗
PubMed1988

Human GAP-43: its deduced amino acid sequence and chromosomal localization in mouse and human.

The growth-associated protein (GAP-43) is considered a crucial component of an effective regenerative response in the nervous system. Its phosphorylation by protein kinase C correlates with long-term potentiation. Sequence analysis of human cDNAs coding for this protein shows that the human GAP-43 gene is highly homologous to the rat gene; this homology extends into the 3'-untranslated region. However, the human protein contains a 10 amino acid insert. Somatic cell hybrids demonstrate localization of the GAP-43 gene to human chromosome 3 and to mouse chromosome 16.

Amino Acid Sequence↗