Adenosine pre- and postsynaptic modulation of glutamate-dependent calcium
activity in hypothalamic neurons.
Obrietan, Karl, Andrei B. Belousov, H.Craig Heller and Anthony N. van den Pol.
Department of Biological Sciences, Stanford University, Stanford CA 94305
and Section of Neurosurgery, Yale University, School of Medicine, New Haven CT
06520.
APStracts 2:0200N, 1995.
SUMMARY AND CONCLUSIONS
1. Within the hypothalamus, adenosine has been reported to influence
temperature regulation, sleep homeostasis, and endocrine secretions. The
effects of adenosine on hypothalamic neurons have not been studied at the
cellular level. Adenosine (5 nM-30 [mu]M) showed no influence on intracellular
Ca 2+ or electrical activity in the presence of glutamate receptor antagonists
D-2-amino-5-phosphonovalerate (AP5) and 6-cyano-7-nitroquinoxaline-2,3-dione
(CNQX); consequently, we examined the role of adenosine in modulating the
activity of glutamate in cultured hypothalamic neurons (n>1700) with fura-2 Ca
2+ digital imaging and whole-cell patch-clamp electrophysiology in the absence
of glutamate receptor block. 2. When glutamate receptors were not blocked,
adenosine (1-30 [mu]M) and the selective adenosine A 1 receptor agonist N 6 -
cyclopentyl adenosine (CPA; 5 nM- 1 [mu]M) caused a large reduction in
intracellular Ca 2+ and electrical activity, suggesting that glutamate
neurotransmission was critical for an effect of adenosine to be detected.
Neuronal Ca 2+ levels were reversibly depressed by CPA (50 nM), with a maximum
depression of 90%, and these effects were blocked by co-administration of the
A 1 receptor antagonist 8-cylopentyl-1,3-dipropylxanthine (DPCPX). 3. Ca 2+
levels in immature neurons prior to the time of synaptogenesis were not
affected by adenosine. Adenosine A 1 receptor activation suppressed glutamate-
mediated Ca 2+ activity in neurons throughout in vitro development from 8 days
to 73 days. 4. Adenosine (1 or 10 [mu]M) caused a hyperpolarization of
membrane potential and a reduction of large postsynaptic potentials arising
from endogenously released glutamate. The administration of low concentrations
of CPA (5 nM) decreased the frequency of glutamate-mediated, neuronally-
synchronized Ca 2+ transients and the frequency of postsynaptic potentials. 5.
To compare the relative effects of adenosine on hypothalamic neurons with
cells from other brain regions, we assayed the effects of CPA on glutamate-
mediated Ca 2+ in hippocampal and cortical cultures. CPA (50 nM) reversibly
depressed glutamate-mediated Ca 2+ rises in hypothalamic neurons by 35%,
compared with 54% in hippocampal neurons and 46% in cortical neurons. 6. If it
does play a functional role, adenosine should be released by hypothalamic
cells. In some neurons the adenosine A 1 receptor antagonists CPT or DPCPX
caused an increase in intracellular Ca 2+ , suggesting that adenosine was
secreted by hypothalamic cells, tonically depressing glutamate-enhanced
neuronal Ca 2+ . 7. To determine if adenosine could exert a postsynaptic
effect, it was co-applied with glutamate agonists in the presence of TTX.
Within subpopulations of hypothalamic neurons, adenosine and CPA either
inhibited (18% of total neurons) or potentiated (6% of total neurons)
responses to glutamate, N-methyl-D-aspartate and kainate by ¯> 20%. 8. In
contrast to the modest effects found in neurons, responses of hypothalamic
astrocytes to the application of glutamate, or the metabotropic glutamate
receptor agonist ( ¯+ )-trans-1-amino-1,3-cyclopentanedicarboxylic acid (t-
ACPD) were strongly potentiated by adenosine (mean= +225%) and CPA. 9.
Together, these findings suggest that adenosine exerts a major presynaptic
effect and minor postsynaptic effect in the modulation of glutamate
neurotransmission in the hypothalamus where it can play a significant role in
blocking a large part of the glutamate-induced Ca 2+ rise. In the absence of
glutamate transmission, adenosine has relatively little effect on either
neuronal intracellular Ca 2+ or on electrical activity.
Received 19 April 1995; accepted in final form 21 June 1995.
APS Manuscript Number J267-5.
Article publication pending J. Neurophysiol.
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 30 July 1995.