Project Details


The function of the 5-hydroxytryptamine3 (5-HT3) receptor is enhanced by
alcohols, but the mechanism(s) of action remains unknown. In addition,
regulation of the 5-HT3 receptor by post-translational modification events,
such as phosphorylation, has not been well characterized. To date, one
subunit of the 5-HT3 receptor has been cloned, and it possesses multiple
consensus sequence sites for protein kinase dependent phosphorylaiton.
Previous work with other ligand-gated ion channels, namely the GABA A and
NMDA receptors, has suggested that protein kinase C may be involved in
mediating these receptors' responsiveness to ethanol. Therefore, it is
reasonable to speculate that kinases may modulate the responsiveness of the
5-HT 3 receptor to alcohols. In the present proposal, functional studies
of the 5-HT3 receptor will be performed using Xenopus laevis oocytes as an
expression system. Two-electrode voltage clamp electrophysiological
recordings of expressed wild-type or mutagenized 5-HT3 receptors will be
made. First, the sensitivity of the 5-HT3 receptor to alcohols will be
measured. Next, the ability of serine/threonine kinases to alter 5-HT3
receptor function will be addressed. To determine which amino acid(s) are
substrates for these kinases, candidate serines in consensus sequences for
protein kinase dependent phosphorylation in the receptor will be mutated to
nonphosphorylatable residues. Kinases which alter 5-HT3 receptor function
will be examined for their ability to change the sensitivity of this
receptor to alcohols. The alternative hypothesis, that tonic kinase
activity underlies the 5-HT3 receptor's sensitivity to alcohols, will be
tested by measuring alcohol modulation of 5-HT3 receptors with mutagenized
consensus sequence sites for phosphorylation. Thus, the ultimate goal of
the proposal is to determine if phosphorylation events alter the function
of the 5-HT3 receptor and its modulation by alcohols. A better
understanding of the regulation of the 5-HT3 receptor is important, given
that this receptor may be involved in mediating the rewarding actions of
drugs of abuse such as ethanol.
Effective start/end date1/05/9530/04/96