- Email: [email protected]
Ethanol and neuronal metabolism
57
K PROTEIN PHOSPHORYLATION ANNETTE Department CT 06510
C. DOLPHIN*, of Pharmacology, (U.S.A.)
SUSAN
IN THE BRAIN
E. GOELZ
Yale
University
and PAUL GREENGARD School
of Medicine,
New
Haven,
Our research activities are currently concerned with discovery and characterization of the protein phosphorylation systems associated with three classes of intracellular messengers, namely cyclic AMP, cyclic GMP and calcium, with the attempt to relate the biochemistry of these phosphorylation systems to functioning of nerve cells, and hopefully with evaluation of the possible role of these systems in various types of neurological and mental illnesses, including alcoholism. We have purified and characterized cyclic AMP-dependent, cyclic GMP-dependent and calcium-dependent protein kinases, and are now engaged in the purification and characterization of specific substrates for these three classes of protein kinases in nervous tissue. In addition, we are attempting to correlate the state of phosphorylation of these specific substrates with various physiological parameters. One substrate protein which we have studied in great detail, referred to as protein I, is a prominent substrate for both a cyclic AMP-dependent as well as a calciumdependent protein kinase in nervous tissue. Protein I appears to be associated primarily with synaptic vesicles and may be involved in the regulation of neurotransmitter release. Another protein which we have studied is a specific substrate for cyclic GMP-dependent protein kinase, and is referred to as the “23K G-substrate”. This substrate appears to be localized primarily in the Purkinje cells of the cerebellum. The availability of this natural substrate protein should help in understanding the role of cyclic GMP in brain function. Studies are in progress to evaluate the effects of alcohol on protein I and on the “23K G-substrate”. Supported by USPHS Grants AA-04183, DA-01627, NS-08440 and a grant from the McKnight Foundation.
MH-17387
and
L ETHANOLANDNEURONALMETABOLISM PAUL MANDEL*, Centre
MARC LEDIG
de Neurochimie
du CNRS,
and JEAN-REM1 11 rue Humann,
M’PARIA 67085
Strasbourg
Ctdex
(France)
The effects of ethanol on nerve cell metabolism were investigated in tissue culture. Attention was focused on the metabolism of alcohol, the activities of membrane-bound enzymes and on superoxide dismutase activity. Clonal cell lines of primary cultures of either pure neurones or pure glial cells were used.
58
A decrease in (Na+,K’)-dependent ATPase activity was observed in clonal cell lines and primary cultures of neurones but not of glial cells. 5’-Nucleotidase activity of isolated neurones and glial cells from chick brain did not change. A slight increase was observed in cultured rat astroblasts over a short period of time. The presence of ethanol in the culture medium produced a decrease of superoxide dismutase activity without significant change in neurones. There was a significant increase of alcohol dehydrogenase activity in cells of a clonal cell line. Most of the changes observed in tissue culture were also found in uiuo. The correlation between findings in vitro and in uiuo will be discussed. 60 NEURONAL MICE PAULA
MEMBRANE
L. HOFFMAN*,
CHARACTERISTICS
MILICA
Department of Physiology West Side Medical Center,
LEVENTAL+
IN ETHANOL-TREATED
and BORIS
and Biophysics, University Chicago, IL 60612 (U.S.A.)
TABAKOFF
of Illinois Medical
Center,
and VA
The neuronal cell membrane has been regarded as an initial site of ethanol action. Adaptive responses to the perturbing effect of ethanol on membrane structure and function may involve changes in lipid composition which alter the physical characteristics of the membrane. Such changes would be expected to influence the function of membrane-bound proteins. In particular, the temperature dependence of the activity of certain membranebound enzymes is related to their microenvironment. Arrhenius plots of the activity of synaptosomal (Na+-K’)ATPase obtained from ethanol-tolerant C57Bl mice revealed that the transition temperature of this enzyme - a parameter strongly influenced by membrane characteristics -was lower than that of enzyme from control animals. Furthermore, while ethanol added in vitro lowered the transition temperature of enzyme from control animals, the enzyme from the mice treated chronically with ethanol was resistant to this effect. The time course of the appearance and disappearance of these changes correlated with that of functional tolerance to ethanol. (Na+-K’)ATPase is an enzyme which spans the entire neuronal membrane. In order to evaluate more precisely the location of membrane changes in response to ethanol, we are examining the effects of acute and chronic ethanol treatment on the temperature dependence of adenylate cyclase, an enzyme localized to the inner leaflet of the neuronal membrane. This enzyme is obtained for our studies from both the caudate. nucleus and cortex, in contrast to the whole brain (Na+-K+)ATPase preparation. Arrhenius plots of basal adenylate cyclase ‘Present address: Belgrade, Yugoslavia.
Institute
for Biological
Research,
University
of Belgrade,
11000
K PROTEIN PHOSPHORYLATION ANNETTE Department CT 06510
C. DOLPHIN*, of Pharmacology, (U.S.A.)
SUSAN
IN THE BRAIN
E. GOELZ
Yale
University
and PAUL GREENGARD School
of Medicine,
New
Haven,
Our research activities are currently concerned with discovery and characterization of the protein phosphorylation systems associated with three classes of intracellular messengers, namely cyclic AMP, cyclic GMP and calcium, with the attempt to relate the biochemistry of these phosphorylation systems to functioning of nerve cells, and hopefully with evaluation of the possible role of these systems in various types of neurological and mental illnesses, including alcoholism. We have purified and characterized cyclic AMP-dependent, cyclic GMP-dependent and calcium-dependent protein kinases, and are now engaged in the purification and characterization of specific substrates for these three classes of protein kinases in nervous tissue. In addition, we are attempting to correlate the state of phosphorylation of these specific substrates with various physiological parameters. One substrate protein which we have studied in great detail, referred to as protein I, is a prominent substrate for both a cyclic AMP-dependent as well as a calciumdependent protein kinase in nervous tissue. Protein I appears to be associated primarily with synaptic vesicles and may be involved in the regulation of neurotransmitter release. Another protein which we have studied is a specific substrate for cyclic GMP-dependent protein kinase, and is referred to as the “23K G-substrate”. This substrate appears to be localized primarily in the Purkinje cells of the cerebellum. The availability of this natural substrate protein should help in understanding the role of cyclic GMP in brain function. Studies are in progress to evaluate the effects of alcohol on protein I and on the “23K G-substrate”. Supported by USPHS Grants AA-04183, DA-01627, NS-08440 and a grant from the McKnight Foundation.
MH-17387
and
L ETHANOLANDNEURONALMETABOLISM PAUL MANDEL*, Centre
MARC LEDIG
de Neurochimie
du CNRS,
and JEAN-REM1 11 rue Humann,
M’PARIA 67085
Strasbourg
Ctdex
(France)
The effects of ethanol on nerve cell metabolism were investigated in tissue culture. Attention was focused on the metabolism of alcohol, the activities of membrane-bound enzymes and on superoxide dismutase activity. Clonal cell lines of primary cultures of either pure neurones or pure glial cells were used.
58
A decrease in (Na+,K’)-dependent ATPase activity was observed in clonal cell lines and primary cultures of neurones but not of glial cells. 5’-Nucleotidase activity of isolated neurones and glial cells from chick brain did not change. A slight increase was observed in cultured rat astroblasts over a short period of time. The presence of ethanol in the culture medium produced a decrease of superoxide dismutase activity without significant change in neurones. There was a significant increase of alcohol dehydrogenase activity in cells of a clonal cell line. Most of the changes observed in tissue culture were also found in uiuo. The correlation between findings in vitro and in uiuo will be discussed. 60 NEURONAL MICE PAULA
MEMBRANE
L. HOFFMAN*,
CHARACTERISTICS
MILICA
Department of Physiology West Side Medical Center,
LEVENTAL+
IN ETHANOL-TREATED
and BORIS
and Biophysics, University Chicago, IL 60612 (U.S.A.)
TABAKOFF
of Illinois Medical
Center,
and VA
The neuronal cell membrane has been regarded as an initial site of ethanol action. Adaptive responses to the perturbing effect of ethanol on membrane structure and function may involve changes in lipid composition which alter the physical characteristics of the membrane. Such changes would be expected to influence the function of membrane-bound proteins. In particular, the temperature dependence of the activity of certain membranebound enzymes is related to their microenvironment. Arrhenius plots of the activity of synaptosomal (Na+-K’)ATPase obtained from ethanol-tolerant C57Bl mice revealed that the transition temperature of this enzyme - a parameter strongly influenced by membrane characteristics -was lower than that of enzyme from control animals. Furthermore, while ethanol added in vitro lowered the transition temperature of enzyme from control animals, the enzyme from the mice treated chronically with ethanol was resistant to this effect. The time course of the appearance and disappearance of these changes correlated with that of functional tolerance to ethanol. (Na+-K’)ATPase is an enzyme which spans the entire neuronal membrane. In order to evaluate more precisely the location of membrane changes in response to ethanol, we are examining the effects of acute and chronic ethanol treatment on the temperature dependence of adenylate cyclase, an enzyme localized to the inner leaflet of the neuronal membrane. This enzyme is obtained for our studies from both the caudate. nucleus and cortex, in contrast to the whole brain (Na+-K+)ATPase preparation. Arrhenius plots of basal adenylate cyclase ‘Present address: Belgrade, Yugoslavia.
Institute
for Biological
Research,
University
of Belgrade,
11000