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CHANGES IN ACTIVITIES OF SODIUM PUMP ISOFORMS AFTER STIMULATION OF CULTURED CEREBRAL NEURONS BY GLUTAMATE. NOBUO INOUE, YOSHIKO ISHIHARA* AND HIDE0 MATSUI* Department of Biochemistry, --L---L Kyorin

University School of Medicine, 6-20-Z Shinkawa, Mitaka, Tokyo 181, Japan. -- Rat cerebral neurons, matured in primary culture, contain isoforms of the Na pump, a weakly

digitalis-sensitive (aI) and a highly digitalis-sensitive (ctI1 and aII1) isoform. Here, the

effects of glutamate on the activities of the Na pump isoforms were examined by measuring 42K+

uptake into neurons. K+ efflux and Na+ influx of the neurons increased during stimulation of the

neurons by glutamate, and subsequently the total Na pump activity increased l&fold. The increase

in the total Na pump activity is due to 3.5-fold increase in the activity of the highly digitalis-

sensitive isoform, a neuron-specific isoform. In contrast, the activity of the weakly sensitive

isoform, an isoform common in various tissues and cells, decreased O&fold after the glutamate

stimulation. These results suggest that the Na pump isoforms are different with physiological significance and that the neuron-specific isoform of the Na pump plays an important role in

recovering the reduced concentration gradients of Nat and Kt In neurons after neuronal excitation.

ONTOGENY OF PHORBOL ESTER RECEPTORS IN THE RAT BRAIN STUDIED BY IN VITRO AUTORADIOGRAPHY. SHOZO KIT0 AND RIE MIYOSHI*, Third Department of Internal Medicine, Hiroshima University School of Medicine, l-Z-3 Kasumi, Minamiku, Hiroshima 734, Japan.

It has been suggested that protein kinase C is a major protein phosphorylation system throughout the developmental process of the brain. In the present study, the ontogeny of phorbol ester receptors, which have been considered to correspond t

9 protein kinase C, in the

rgt brain was examined through in vitro autoradiography with H-phorbol ( H-PDBu).

13,13-dibutyrate From binding experiments using cryostat sections, ft was found that H-PDBu had a

single high affinity binding site whose Kd value was 11.7 nM. H-PDBu binding sites were highly concentrated in the olfactory tubercle, striatum, cerebral cortex, amygdala, hippocampus, yubstantia nigra, H-PDBu binding

molecular layer of the cerebellum3and so on. The developmental pattern of sites varied with brain region. H-PDBu binding sites in the amygdala,

thalamus, stratum pyramidale of CA1 of the hippocampus, dentate gyrus, superior colliculus, substantia nigra, interpeduncular nucleus, and cerebellar molecular layer were postnatally increased to adult levels and after that remained constant. On the other hand, in the stratum oriens an stratum radiatum of CA1 of the hippocampus, and in the lateral and medial geniculate bodies,

$ H-PDBu binding sites reached peaks at 21 or 28 days of postnatal age and afteg that

they declined to adult levels. The cerebellar granule cell layer showed a low level of H-PDBu binding sites throughout all the ontogenetic stages. A distinct ontogenetic pattern of phorbol ester receptors in various regions of the brain may reflect a role of protein kinase C in the neural development of each discrete area.

QUANTITATIVE AUTORADIOGRAPHICAL LOCALIZATION OF PHORBOL ESTER BINDING SITES IN MONKEY BRAIN. YUMIKO WATANABElr2, PER-GORAN GILLBERG3, SHOGO TSUBOKURAl, AND YASUYOSHI WATANABEI. IDepartment of Neuroscience, Osaka Bioscience Institute, Osaka 565, 2Suntory Institute for Biomedical Research, Osaka 618, and 3University of Uppsala, Sweden.

Japan,

Quantitative autoradiographical localization of 3H phorbol 12,13-dibutyrate (PDBu) binding sites was investigated using Macaca fuscata hemisphere coronal sections (50-pm thickness). The PDBu binding activity was high (500-1500 fmol/mg tissue) in the CA1 and dentate gyrus of the hippocampus, medial preoptic area, substantia nigra pars diffusa, molecular layer of the cerebellum, and substantia gelatinosa and lamina X of the spinal cord, and was moderate (200-500 fmol/mq tissue) in amygdaloid N., caudoputamen, N. accumbens, claustrum, septum, subiculum of the hippocampus, medial geniculate N., and central gray of midbrain. In the cerebral cortex, layer I-II showed the highest PDBu binding (454 fmol/mg tissue), followed by layer II-VI (253 fmol/mg tissue), whereas layer IV in the visual cortex exhibited low levels. These results may provide basic data for understanding the role of protein kinase C (specific PDBu binding site) in higher brain function through the use of positron emission tomography and neurophysiological tasks on primates.