NICOTINE MEDIATED SODIUM FLUX IN PRIMARY CULTURES

Eighty percent of smokers who quit relapse within a year. Although there
are many reasons for relapse, the active ingredient in cigarettes,
nicotine, has properties of an addictive substance. All of the actions of
nicotine are not known, but nicotine is known to act as receptor sites in
nervous tissues. Its action at acetylcholine receptor sites may be related
to its addictive properties. The nicotinic acetylcholine receptors in mammalian brain have not been
well-characterized. Nicotine binds with high affinity (Kd 1-10 nM) to at
least one population of brain nicotinic receptors. The high-affinity
binding to brain tissue is a paradox. Electrophysiological studies suggest
that micromolar concentrations of nicotine are necessary for receptor
activation. It has been suggested that the brain receptors are desensitized
during the assay procedure (in which 10-20 nM concentrations of nicotine
are used), indicating a 1000-fold difference between the K-activation (K-
act) and K-desensitization (K-des) of nicotinic receptors in mammalian
brain. Previous work in our laboratory indicates that the K-act and K-des of PC12
cells, which possess a well-characterized neuronal receptor, are 7.7 X
10(5) M and 8.2 X 10(5) M, respectively. On the basis of these data, we
question whether the K-des and K-act in brain tissue would differ by 3
orders of magnitude. This question cannot be resolved using brain tissue
in vivo. We therefore propose to study this question in primary neuronal
brain cultures using 22Na+ influx. 22Na+ influx has not previously been demonstrated in primary neuronal
cultures derived from mammalian brain. Therefore, the goal of this research
is to demonstrate nicotinic receptor mediated 22Na" influx in mammalian
brain culture. Experiments using 22Na+ influx and nicotine and cytisine as
agonists will be performed to determine the K-act and K-des of the
receptors. Influx will be also carried out in the presence of nicotinic
antagonists. These experiments will serve as the basis for future inves-
tigations of the physiological properties of brain nicotinic receptors.