Although disulfiram used as a pharmacological agent in the treatment of alcoholism is reported to act on both peripheral and
central nervous systems with several adverse effects, the ncurotoxic property of the drug has not been properly elucidated. We
observed that the chronic administration of the drug to rats significantly inhibited synaptosomal (Na+,K+)-ATPase and basal
Mg2+-ATPase activities. Further, the uptake of y-aminobutyric acid and e-glutamate which rely on the energy provided by this
system was depleted following chronic drug administration. Similar findings were observed when the isolated synaptosomes were
treated with the drug in an in vitro system. Further, treatment of synaptosomes with ouabain, a known inhibitor of
(Na+,K +)-ATPase resulted in significant depletion of 3H-GABA and e-[3H]glutamate uptake into synaptosomes indicating the
importance of the enzyme in the uptake mechanism. However, diethyldithiocarbamate, a major metabolite of disuifiram did not
elicit any change in either the enzyme activity or the uptake of these neurotransmitters. On the basis of these evidences, we
suggest that the chronic disulfiram administration attenuated the neurotransmitter uptake mechanism and resulted in higher
cxtracellular concentration of glutamate that could lead to glutamate-induced neurotoxicity.


The uptake of excitatory neurotransmitter L-gluta-
mate (L-GIu) and inhibitory neurotransmitter y-amino

butyric acid (GABA) into synaptosomes exclusively rely
on the energy provided by (Na +,K รท)-ATPase system of
the brain. Indeed the neurotransmission by amino acids
is terminated by high affinity uptake of the transmitters
in neurons and glial cells. In fact it was shown that an
impairment of the Na+-dependent glutamate uptake
capacity, which occurs during energy failure of the
brain such as ischemia and anoxia (Sanchez-Prieto and
Gonzalez, 1988; Silverstein et al., 1986), results in
excess amounts of extracellular glutamate. Further
C1–dependent glutamate uptake was observed in
synaptic membranes and glial cells (Cho and
Bannai,1984; Zaczek et al., 1987), and the treatment of
(Na+,K+)-ATPase with ouabain affected Cl–dependent uptake in synaptic membranes (Koyama et al.,
1993). More recently a carrier-mediated release and

uptake of GABA and glutamate has also been re-
ported (see review by Levi and Raiteri, 1993). Over the

last few years much interest has been focussed upon
neurotoxic properties of neuroexcitatory amino acid
glutamate. Although the concept of an excitatory

mechanism as the basis of toxic effects is still contro-
versial, there can be no doubt to date as to potential of glutamate as a neurotoxin.

Among the compounds that augment neurotoxicity,
we discovered tetraethylthiuram disulfide (disulfiram)
an extensively used pharmacological agent for the
treatment of alcoholism. Although, the disulfiram has
been known to cause neurotoxicity (Laplane et al.,
1992; Simonian et al., 1992), the precise mechanism of

its neuropathological role is yet to be elucidated. How-
ever, the electrophysiological findings suggest that

disulfiram induces degeneration of the axon and basal
ganglia. These neuropathies occur even in the absence
of disulfiram ethanol reaction suggesting that factors other than the inhibition of aldehyde dehydrogenase and dopamine fl-hydroxylase are involved in the neuro- toxic effects of disulfiram. Experimental studies have shown that serotonergic (Fukumori et al., 1980; Nagen-
dra et al., 1993), dopaminergic (Molinengo et al., 1991), noradrenergic and cholinergic (Nilsson et al., 1987) functions are affected by disulfiram. It has been re-
ported that disulfiram modulates Mg2+-ATPase-de – pendent uptake of monoamines in chromaffin granular
membranes (Schlichter et al., 1975). Disulfiram also
interacts with plasma membranes affecting its integrity, and may alter the native structure of proteins by form-
ing internal S-S bonds (Brien and Loomis, 1985; Lauri-
alt and O’Brien, 1991). Thus, we expect disulfiram to

follow the same cellular mechanism that alters the
membrane integrity and affects membrane transport
eventually leading to the neurotoxic effect. We assume

that the development of cell injury/death is a conse-
quence of membrane perturbation and transport mech-
anism caused by disulfiram.

In the present study we have examined the effects of
brief and chronic treatments with disulfiram on
(Na+,K+)-ATPase activity and the uptake of GABA
and L-GIu into the brain synaptosomes.

Materials and methods

L-[3,4-aH]glutamate (specific activity 5.19 Ci/mmol)
and y-[2,3-3H]aminobutyric acid (specific activity 94.2
Ci/mmol) were purchased from DuPont-New England

Nuclear (USA). All other chemicals used were of ana-
lytical grade including disulfiram, diethyldithio-
carbamate and ouabain obtained from Sigma Chemical

Co. (St.Louis, MO, USA).