CA2+/CALMODULIN-KINASE II--BRAIN SUBSTRATES

The objective of these studies is to identify physiological
functions of Ca++/ Calmodulin-dependent protein kinase II (CaM-
kinase II) in neural tissues. CaM-kinase II is of particular
interest in this context since 1) it constitutes about 1% of brain
protein, 2) it constitutes about 50% of the postsynaptic density
(PSD) protein, and 3) it undergoes a unique autophosphorylation
that converts it to a Ca++ -independent form. Our studies will
utilize a combination of biochemical and electrophysiological
approaches to investigate physiological functions of the cytosolic
and membrane-associated brain CaM-kinase II. Potential targets of cytosolic CaM-kinase II that will be
investigated include tyrosine hydroxylase, the rate-limiting enzyme
in catecholamine and several other regulatory CaM-binding proteins.
With reference to tyrosine hydroxylase, our goals will be to
establish its in vivo phosphorylation by CaM-kinase II and the
regulatory role of an activator protein that is specific for
tyrosine hydroxylase that has been phosphorylated by CaM-kinase
II. These studies will utilize pinocytotic introduction of
antibodies against CaM-kinase II or the activator protein into PC12
cells. A number of known CaM-binding proteins will be screened for
specific phosphorylation by the Ca -independent form of CaM-kinase
II. We are especially interested in regulatory phosphorylation
sites which are blocked when Ca /CaM is bound to the CaM-binding
proteins. A major focus of these studies will be the CaM-kinase II localized
in the PSD. Of special interest is the potential role of this
kinase in regulating certain ion channels, specifically, the NMDA-
receptor/ion channel and the dihydropyridine-sensitive Ca channel.
These studies will combine radioligand binding analyses and patch-
clamp studies. We are particularly interested in potential
regulation of these ion channels by protein phosphorylation and by
GTP-binding proteins. The NMDA channel is of special relevance due
to its likely involvement in synaptic plasticity such as long-term
potentiation and kindling, and perhaps in epilepsy.