摘要:

AbstractMuscle (body wall, buccal mass, heart) and neural tissue of the marine molluscAplysia californicawas analysed for calcium‐binding proteins using transblot45Ca overlay, Western blotting and two‐dimensional polyacrylamide gel electrophoresis, and several low molecular weight calcium‐binding proteins were identified. Our results show thatAplysiamuscle contains an abundant protein with aMrof ∼20 000 with strong45Ca2+‐binding ability and cross‐reactivity to antibodies against the sarcoplasmic calcium‐binding protein isoform II (SCP II) fromAmphioxus.Immunocytochemical studies revealed that isoforms of SCP are distributed in a tissue‐specific manner, SCP II‐like protein is exclusively present in muscle fibres closely associated with the contractile machinery, whereas the isoform I (SCP I‐like protein) is exclusively present in a subset of neurons, suggesting a function in their calcium regulation. In addition, a novel45Ca2+‐binding protein ofMr43 000, pl 4.7, was found in muscle and in neurons. A third protein ofMr40 000, pl 4.8, cross‐reacts with anti‐parvalbumin and anti

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00520.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractA depolarization‐induced, slowly decaying inward current was examined in slice‐cultured CA3 pyramidal cells by voltage‐clamp techniques and microfluorometric measurements of cytosolic free Ca2+concentration ([Ca2+]i). Action potentials elicited by intracellular injection of short‐lasting (50 – 100 ms) depolarizing current pulses were followed by a slowly decaying afterhyperpolarization (AHP). After switching to voltage‐clamp mode, short‐lasting (50 – 100 ms) depolarizing voltage jumps from –60 mV to between –30 and 0 mV induced a slowly decaying outward aftercurrent (IAHP) which was depressed by bath application of muscarine (0.5 μM). In the presence of muscarine, the same depolarizations induced a slowly decaying afterdepolarization (ADP) or inward aftercurrent (IADP)in voltage‐clamp mode. This current was also induced in the presence of trans(±)‐1‐aminc‐1,3‐cyclopenta‐nedicarboxylic acid (t‐ACPD, 5 μM), an agonist of metabotropic glutamate receptors, but not in the presence of noradrenalin (5 μM), while both of these agonists depressed IAHP. IADPwas depressed by reducing the external Ca2+concentration from 3.8 to 0.5 mM, by external Co2+(1 mM) and by external Cd2+(10 – 100 μM). Combined voltage‐clamp recordings and microfluorometric measurements of [Ca2+]iusing the Ca2+indicator fura‐2 revealed that the amplitude of IADPwas correlated with the amplitude of depolarization‐induced Ca2+influx, IADPwas absent at membrane potentials<–90 mV, and reached maximal amplitudes at ∼–55 mV. Raising the extracellular K+concentration from 2.7 to 13.5 mM increased the amplitude of IADPand resulted in a positively directed shift of the apparent reversal potential of IADP. When the external Na+concentration was reduced from 157 to 33 or 18 mM the current reversed at more negative potentials and was reduced to 40 and 21%, respectively, of control amplitude. Lowering the external Cl‐concentration from 159 to 20 mM did not affect IADP. We conclude that IADPmost likely represents a Ca2+‐activated cation current, rather than a Ca

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00521.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractSeveral lines of evidence suggest that excitatory synaptic transmission contributes to the maturation of precise neuronal connections. In the present study we determined whether the specific innervation pattern of single inhibitory arborizations was dependent upon neuronal activity during development. A homogeneous group of glycinergic inhibitory neurons in the central auditory system, the medial nucleus of the trapezoid body (MNTB), was functionally denervated in neonatal gerbils. The anatomical specificity of single MNTB terminal arborizations was subsequently measured along the tonotopic axis of a postsynaptic target, the lateral superior olive. Here we demonstrate that inhibitory terminal boutons spread a significantly greater distance along the frequency axis of the postsynaptic target following functional denervation. Although total arbor length remained unchanged, there was a significant increase in the number of branch points, suggestingde novosprouting. The results indicate that normal inhibitory synaptic activity contributes to the developmental refinement of specific neuronal connections.

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00522.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractIn order to investigate signalling pathways involved in the control of granule cell differentiation, survival and other functions by depolarization or activation of NMDA receptors we have characterized protein phosphorylation in cerebellar granule cells. Cultures of cerebellar granule cells were incubated with32P orthophosphate and then challenged with NMDA, K+or the Ca2+ionophore ionomycin, agents which raise [Ca2+]iand stimulate differentiation and survival. Upon separation of labelled phosphoproteins by two‐dimensional gel electrophoresis three differences were found in response to all of these agents. These were an increase in acidity of two phosphoproteins of 87 and 48 kDa (p87 and p48) and increased32P‐incorporation into a phosphoprotein of 120 kDa (p120). Treatment with PMA which stimulates neurite outgrowth but not survival affected p87 (increased its acidity) but not p48. The acidic shift of p87, therefore, is not sufficient to stimulate granule cell survival. The identification of p87 as the actin‐binding MARCKS protein and the demonstration of its presence in neurites and growth cones of granule cells suggests that it may be involved in NMDA‐stimulated neurite outgrowth. The phosphoproteins p120 and p48 may potentially be involved in events linking the rise in [Ca2+]ito increased granule cell survival or other aspects of granule cell differen

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00523.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractHorizontal cells, which are second‐order neurons of the vertebrate retina, exhibit synaptic plasticity governed by light and dark adaptation. We have investigated the alterations in the protein phosphorylation patterns of isolated carp (Cyprinus carpio) horizontal cells in relation to their state of light adaptation by using anin vitrophosphorylation assay and compared the resulting data with protein synthesis patterns of the whole retina. Phosphoproteins and [35S]methionine‐labelled proteins were analysed by one‐ and two‐dimensional gel electrophoresis followed by autoradiography. The state of light adaptation significantly affected thein vitrophosphorylation of horizontal cell proteins with molecular weights of 68, 56/58, 47, 28 and 15 kDa, but had no effect on the protein synthesis of retinal proteins. In the light the most prominent increase of32P incorporation was observed in the 47 kDa protein. The biochemical properties of this protein closely resembled those of the growth‐associated GAP‐48, found in the fish retina. In addition, the phosphorylation of horizontal cell homogenates in the presence of protein kinase activators such as cyclic AMP, calcium, calmodulin and phospholipids revealed that horizontal cells of the fish retina contain cyclic AMP‐, calcium/calmodulin‐ and calcium/phospholipid‐dependent protein kinase activity resulting in the phosphorylation of several horizontal cell proteins, including the phosphoproteins which were affected by the state of

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00524.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractIn vivo, the surface glycoprotein Schwann cell myelin protein (SMP) is expressed in the quail peripheral nervous system exclusively by Schwann cells. It is not detectable at any developmental stage either in enteric glia or in ganglionic satellite cells. We demonstrate here that the satellite glial cells of the dorsal root ganglia start to express SMP on their surface when they are dissociated into single cells and cultivatedin vitro.Activation of SMP synthesis is a rapid event observed in mass cultures of dorsal root ganglia dissociated cells as soon as 4 h after the onset of the culture. Confocal microscope analysis revealed that satellite cells may acquire the Schwann cell marker when still in close contact with the neuronal soma. Clonal cultures of satellite cells from E8 dorsal root ganglia demonstrated that the progeny of these SMP‐negative cells steadily express SMP. This, together with similar results previously obtained with enteric glia, suggests that the SMP‐positive phenotype is a constitutive trait of the peripheral glial cell lineage which is inhibited in satellite cellsin vivoby the microenvironment prevailing in the peripheral nervous system gang

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00525.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractThe neurotrophin gene family includes four structurally related proteins with neurotrophic activities. Two of them, nerve growth factor and brain‐derived neurotrophic factor (BDNF), have been studied in detail and information has recently emerged on the expression and function of the third member, neurotrophin‐3. In contrast, little information is available on neurotrophin‐4 (NT‐4), the most recently isolated member of this family. In this report we have used a sensitive RNAase protection assay to analyse the developmental expression of NT‐4 mRNA in the rat brain and in 12 different rat peripheral organs. In heart, liver and muscle plus skin NT‐4 mRNA levels were maximal at embryonic day (E) E13 (the earliest time point tested), with reduced levels at later times of development. In lung, kidney and thymus similar levels were seen from E13 to postnatal day (P) 1, with reduced levels in the adult. In testis, ovary and salivary gland NT‐4 mRNA was detected at E16 with a peak shortly after birth. During brain development, NT‐4 mRNA was maximal at E13 followed by a decrease around birth, after which the level increased. The postnatal increase of NT‐4 mRNA was also seen in cerebral cortex and brain stem analysed separately, while in the hippocampus similar levels were found from P1 to adulthood. NT‐4 mRNA was detected in all ten adult rat brain regions analysed with only small regional variations, being highest in pons–medulla, hypothalamus, thalamus and cerebellum. Adult rat thymus, thyroid, muscle, lung and ovary contained higher levels of NT‐4 mRNA than brain, while all other adult tissues showed similar or lower levels than in the brain. The highest level of NT‐4 mRNA overall was found in P1 testis. For comparison, BDNF mRNA was analysed in the same tissues. The expression of BDNF mRNA was in many cases different from that of NT‐4 mRNA. The peak of NT‐4 mRNA expression in several of the peripheral tissues coincided with the peak of naturally occurring neuronal cell death in peripheral ganglia. This is consistent with the possibility that NT‐4 acts as a target‐derived trophic factorin vivo.The widespread and increased expression of NT‐4 mRNA during postnatal brain development could reflect a trophic role of NT‐4 for central nervous system neurons. However, non‐neuronal functions of NT‐4 are also possible, particularly in male and female reproductive tissues, where the NT‐4 protein could function

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00526.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractIn order to identify markers of the two neuronal populations which are successively involved in forebrain ontogenesis, we performed a differential screening of a murine cDNA library with two radiolabelled probes corresponding to striatal mRNAs extracted at embryonic day (E) 17 and E20, i.e. before and after the invasion of the striatum by the late‐born matrix neurons. One of the selected clones, the 3.1 cDNA, corresponds to a very abundant embryonic neuronal transcript enriched in the germinal zones at E17 and in superficial cortical layers and striatum at E20, suggesting that it is expressed mainly in neurons belonging to a late migration wave. During adulthood, it persists at a high level in the granular layer of the cerebellum, the hippocampus and the olfactory bulb, which are the sites of postnatal neurogenesis and intense synaptic plasticity. This 2000 base RNA is enriched in polysomal fractions and encodes a small putative 68 amino acid polypeptide which is conserved in mouse and ma

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00527.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractTo characterize regulatory DNA sequences involved in oligodendroglial expression of myelin basic protein (MBP), transgenic mice carrying a 256 bp fragment of the mouseMBPpromoter fused to anEscherichia coli lacZgene were generated. Of four transgenic families, two (lines 2 and 4) expressed β‐galactosidase activity in the nervous system but not in most other tissues. Histochemical and immunohistochemical analysis of adult brain from these two lines showed oligodendroglial‐specific expression of the transgene. In line 2, only a small proportion of oligodendrocytes expressed the transgene, and in labelled cells the product of the enzymatic reaction with β‐galactosidase was confined to a small round vesicle in the vicinity of the nucleus. In contrast, in tissue sections from line 4 adult brain and spinal cord β‐galactosidase activity was much more intense and at least 80 – 90% of oligodendrocytes expressed the transgene. Detection of theMBP‐lacZtranscript byin situhybridization showed that the transgene mRNA was confined to the oligodendrocyte cell body. These results suggest thatcis‐acting regulatory elements, specifying oligodendrocytes identity, are located within 256 bp upstream

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00528.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

AbstractThe expression of tissue‐ and urokinase‐type plasminogen activators has been studied in developing cerebellum, hippocampus, cerebral cortex, olfactory bulb and olfactory mucosa of the rat byin situhybridization. All identifiable neurons express urokinase mRNA from an early stage in their development, and this expression appears to coincide with the onset of axogenesis. For cerebellar granule cells, both axonal growth and urokinase expression are initiated before they migrate from the external granule layer; for the majority of neocortical neurons, however, both processes are commenced after the cells have migrated to the cortical plate. Neurons continue to express this protease in the adult. The large projection neurons exhibit the highest levels of message, the smaller interneurons having much lower levels except for hippocampal granule cells, which have notably high levels of expression. Glial cells generally do not express urokinase message, except for transient expression by oligodendrocytes in developing fibre tracts during the period of myelination. Thus for both neurons and oligodendrocytes, the onset of urokinase‐type plasminogen activator expression coincides with their initiation of major process outgrowth, although neurons maintain this expression in the adult, possibly to retain a degree of synaptic plasticity. In contrast, although high levels of message for the related protease, tissue plasminogen activator, are found in the embryonic floor plate, in postnatal brain it is abundantly expressed only by ventricular ependymal cells and by cells in connective tissue surrounding the olfactory

ISSN:0953-816X    

DOI:10.1111/j.1460-9568.1993.tb00529.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY