摘要:

Abstract:The uptake of calcium was examined in primary cultures of pure neurons and of glial cells from dissociated hemispheres of chick embryo brain. Neuronal cultures took up calcium at a rate of 2.0 nmol per min per mg cell protein at medium concentrations of 1.2 mM‐Ca2+and 5.4 mM‐K+. The rate of calcium entry into neurons was increased 2.7‐fold by elevating medium potassium to 60 MM. The effect of high external potassium was to increase the Vmaxvalue for calcium transport from 5.5 to 13 nmol per min per mg; the Michaelis constant for calcium, 1.2 mM, was unchanged. The potassium‐dependent component of calcium entry into the neuronal cultures was eliminated by addition of 0.1 mM‐D‐600 (a verapamil derivative) or by 1 mM‐CoCl2, but 0.5 μM‐tet‐rodotoxin had no significant effect. When choline replaced potassium in uptake medium no change in calcium transport was detected in neurons, nor was the entry of calcium increased when choline replaced sodium. Glial cultures took up calcium at 20% of the basal rate for neuronal cultures on a weight‐of‐protein basis. Uptake was not increased by potassium; during depolarization by potassium the calcium transport activity of glia was less than 10% that of neurons. It was concluded that cultured neurons contain a depolarization‐sensitive, calcium‐specific channel. A similar calcium transport activity was not detect

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02380.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:The endogenous level of cyclic AMP in incubated synaptosomes from cerebral cortex of guinea pigs was investigated after the addition of various agents to the incubation medium. It appeared that the synaptosomal suspension already contained exogenous adenosine. Preincubation with theophyl‐line or with adenosine deaminase (ADase) decreased both the exogenous level of adenosine and the intrasynaptosomal level of cyclic AMP. The level of cyclic AMP was reincreased by the addition of adenosine agonists, especially 2‐chloroadenosine. This increase was antagonized by deoxyadenosine and was not inhibited by dipyridamole. These results suggest that the adenosine derivatives in the synaptic cleft regulate the level of cyclic AMP in nerve terminals through adenosine receptor on the presynaptic membrane. ADP, ATP, dopamine, and histamine also stimulate the formation of cyclic AMP in the ADase‐treated synapto

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02381.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:Tyrosine hydroxylase, the rate‐limiting enzyme in catecholamine biosynthesis, catalyzes the conversion of tyrosine to DOPA. Cyclic AMP‐dependent protein phosphorylation conditions alter tyrosine hydroxylase activity in rat striatal homogenates. In agreement with other laboratories, we find that short‐term pre‐incubation (3 min) of extracts under phosphorylating conditions (Mg. ATP, cAMP) increases enzyme activity two‐ to tenfold over control as measured during a subsequent 15‐min assay. We now report that pre‐incubation under phosphorylating conditions for longer periods (30 min) results in a loss of activity to levels equal to or below that of the control enzyme. Addition of purified bovine brain protein kinase catalytic subunit and Mg. ATP enhances activation and increases the rate of inactivation. To demonstrate that inactivation is not associated with proteolytic degradation or irreversible de‐naturation, the inactivated form of the enzyme can be reactivated. The protein kinase inhibitor protein decreases the activation process and prevents inactivation of the enzyme to below control values. The sedimentation coefficient is not changed by phosphorylation conditions (S = 8.8 ± 0.1). Although the apparentKm, of the enzyme for the 6‐methyltetrahydropterine (6‐MPH4) cofactor is reduced (0.86 mM, control; 0.32 MM, activated), it is also reduced in the inactivated form (0.38 mM). TheKi, for dopamine is increased from 4.5 μM for the control to 28 μM for the activated enzyme, whereas the inactivated form of the enzyme exhibits aKiof 10 μM. Removal of catecholamines by gel filtration fails to alter activity and the apparent cofactorKm. Moreover, both the activated and the inactivated states persist following gel filtration. It therefore appears that the activation‐inactivation process is not mediated solely by the modulation of enzyme feedback inhibition or changes in theKm, for 6‐MPH4. We also describe a coupled decarboxylase assay in which labeled dopamine is resolved from the precursors tyrosine and DOPA by low‐

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02382.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:Circulating antibody to the bovine white matter proteolipid apoprotein was detected in rabbits 1 month after a single injection of the water‐soluble form of the apoprotein. By double immunodiffusion, the antiserum reacted specifically with the delipidated proteolipid apoprotein and the crude proteolipid fraction containing complex lipids; after exposure of the proteolipid apoprotein to sodium dodecyl sulfate (SDS), no reactivity was observed. The antiserum did not react with other myelin components, i.e., basic protein, cerebroside or GM1ganglioside, nor was there reactivity with non‐neural proteolipids. The anti‐apoprotein antibody was purified by affinity chromatography. The antibody‐antigen interaction is apparently very hydrophobic, since elution of the antibody from the affinity column requires buffer containing 0.5% Triton X‐100

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02383.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:Antiserum to the catalytic subunit of goldfish brain (Na+,K+)‐ATPase has been employed at the electron microscopic level by means of the peroxidase‐antiperoxidase immunohistochemical method. In optic nerve, an‐tigenic sites are restricted to the nodes of Ranvier. No reaction product is detected in underlying internodal neurolemma. Outgrowing neurites for cultured retinal explants devoid of glial ensheathment exhibit a continuous distribution of the enzyme subunit. Antibodies against eel electroplax (Na+, K+)‐ATPase cross‐react with the goldfish brain enzyme and show a similar immunocytochemical distributio

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02384.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:Synaptosomes isolated on isosmotic Ficoll density gradients are an effective model for some aspects of neuronal function. They maintain metabolic energy levels ([ATP]/[ADP][Pi1) and transplasma membrane electrical potentials very similar to those of neurons in the intact brain. The concentration of K+in the external medium (K+‐sensitive electrode), O2uptake, and cytochromecreduction (550 nm minus 540 nm) were simultaneously monitored in synaptosomal suspensions. Oxidative metabolism is the primary source of intrasynaptosomal ATP and at pH 7.4 anaerobiosis results in K+leakage at 4.5 ± 0.8 nmol/min/mg protein with glucose as substrate and 10.7 T 1.9 nmol/min/mg protein with lactate plus pyruvate (10:1) as substrate. Reintroduction of oxygen initiates complete (ouabain‐sensitive) reuptake of K. at initial rates of 35.4 ± 3.2 nmol/min/mg protein and 18 ± 1.7 nmol K‐/min/mg protein, respectively. The rates of K+leakage and reuptake fall when the pH is lowered from 7.4 to 6.0 but recover fully if the pH is raised to the original value. The rates of K1release and uptake decrease when the Na‐concentration in the medium is decreased and increase when the Ca2‐concentration is decreased. The intrasynaptosomal [K+] under aerobic conditions was 77.3 ± 3 MM and the calculated K+diffusion potential was ‐72 mV. Anaerobic incubation of the synaptosomes for up to 20 min and at pH values from 7.4 to 6.0 did not produce irreversible impairment of any of the measured variables. These results suggest that permanent loss of brain function following prolonged hypoxia and ischemia is not due to irreversible damage to the synapses with respect to these parameters but rather to impairment of some other neu

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02385.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:The effects of gestational date (13, 14, 15, and 17 days) of administration of methylazoxymethanol acetate (20 mg/kg) on the cortical synaptic chemistry and morphology of the rat has been examined in adult offspring. Treatment at 13 days of gestation (DG) resulted in cortical hypoplasia that affected primarily the deep layers whereas treatment at 14 and 15 days gestation caused a severe hypoplasia of cortical neurons above layer V and a 66 to 77% reduction in cortical mass. The 17‐DG treatment caused only a 20% reduction in cortical weight with effects apparent only in the superficial layers. At no treatment date did the specific activity of glutamate decarboxylase differ significantly from control. In contrast, presynaptic markers for noradrenergic (tyrosine hydroxylase and norepinephrine) and for serotonergic (serotonin) terminals were increased in concentration in direct proportion to the degree of cortical hypoplasia. The specific activity of choline acetyltransferase was significantly increased at all treatment dates but total activity per cortical slab was significantly reduced on treatment days 13

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02386.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:Themandpisomers of hydroxyphenylacetic acid have been identified and quantitated in whole rat brain and in several regions using a capillary column high resolution gas chromatography–mass spectrometry procedure. Their concentrations were: form‐hydroxyphenylacetic acid (mean ± S.E., number of determinations in parentheses)—whole brain, 2.3 ± 0.3 ng/g (7); hypothalamus, 1.2 ± 0.3 ng/g (5); caudate nucleus, 5.5 ± 0.6 ng/g (5); brain stem, 1.8 ± 0.1 ngig (5); cerebellum, 1.2 ± 0.1 ng/g (5) and the “rest,” 1.7 ± 0.1 ng/g (5); and forp‐hydroxyphenylacetic acid–whole brain, 10.6 ± 0.7 ng/g (7); hypothalamus, 4.5 ± 0.1 ng/g (4); caudate nucleus, 28.3 ±1.6 ng/g (5); brain stem, 8.6 ± 0.6 ng/g (5); cerebellum, 8.1 ± 0.4 ng/g (9, and the “rest,” 5.3 ± 0.5 ng/g (5). This heterogeneous distribution parallels closely that exhibited by their respective pre

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02387.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:Glutaminc synthetase activity was investigated in developing primary astroglial cultures established from newborn mouse cerebral hemispheres. Between the 2nd and 4th week of culture there was little change in activity under our standard culturing conditions; however, when hydrocortiwne (10 μM) was added to the cultures for 48 h, the enzyme activity increased two‐ to fourfold, depending upon the age of the culture, with maximum response in 2‐week‐old cultures. The addition of dibutyryl cyclic AMP (dRcAMP) to the culture medium caused morphological differentiation of the astroglial cells but eliminated the response of the cells to hydrocortisone. Culturing in elevated serum levels, which delays morphological differentiation and inhibits astroglial cytodifferentiation after exposure to dBcAMP, shifted the time of maximal response to hydrocortisone from 2 to 3 weeks and prevented the abolishment of glutamine synthetase induction by dBcAMP. The induction of glutamine synthetase by hydrocortisone was prevented by actinomycin D (0.5 μg/ml), indicating its dependence upon RNA and protein synthesis. The present work thus confirms reports in the literature that hydrocortisone induces glutamine synthetase in neural tissues, but differs from the findings of Moscona and co‐workers in the chick retina that intact tissues are required for the induction

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02388.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY

摘要:

Abstract:Rat and mouse CNS neurofilament proteins (NFPs) were characterized and compared, in terms of electrophoretic properties on polyacrylamide gels and by peptide mapping, with one another and with other co‐purifying lower‐molecular‐weight CNS proteins, including α and β tubulin. NFPs were partially purified by modification of the axon flotation procedure of Norton and co‐workers and were demyelinated with Triton X‐100. On one‐dimensional SDS polyacrylamide gels the molecular weights of the triad of NFPs from both rat and mouse were approximately 200,000, 140,000, and 70,000. Prominent lower‐molecular‐weight proteins (63,000‐16,000) as well as minor amounts of tubulin and actin were observed after gel electrophoresis. On two‐dimensional gels (isoelectric focusing followed by SDS gel electrophoresis) each of the NFPs appeared to be composed of more than one component and the corresponding NFPs from rat and mouse had similar isoelectric points. Gel electrophoresis peptide mapping usingStaphylococcus aureusV8 protease indicated the following: (1) the triad of NFPs of different sizes have different peptide maps; (2) α and β tubulin have nonidentical digestion products, which are dissimilar to those of the NFPs; (3) other proteins that co‐purify by the axon flotation procedure also have nonidentical peptide maps; and (4) the corresponding NFPs from rat and mouse have similar peptide maps. The co‐purifying proteins examined in detail (63,000–49,000) do not appear to be derived by proteolytic cleavage of NFPs and may represent ot

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1981.tb02389.x

出版商:Blackwell Publishing Ltd    

年代:1981

数据来源: WILEY