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VOL. 76 (1), 3-22, 2010 CROSS-TALK BETWEEN GLUTAMATE AND NUCLEOTIDE...

Once demonstrated that glutamate modulates nucleotide
receptors signalling in cerebellar granule neurons, the main question
was to understand the biochemical components involved in their
interaction. Several evidences suggested that CaMKII could
participate in the cross-talk process. Thus, P2X2, P2X3, P2X4, P2X7
and P2Y1, which distribution in somatic regions from granule
neurons have been previously shown (4, 6), posses sequences
susceptible to be phosphorylated by CaMKII. Furthermore, a large
number of proteins have been described as being associated with
P2X7 (32). One of these proteins is a-actinin, which also interacts
with CaMKII and may be a way to locate CaMKII near its substrates
(33). Concerning to metabotropic P2Y receptor, a previous report
has shown that endocytosis of the P2Y1 receptor is controlled by
the activity of CaMKII (25). Finally, CaMKII is phosphorylated
in response to glutamate receptor activation. Based on these data, it
is probably that CaMKII was involved in the cross-talk mechanism
between glutamate and nucleotide receptors. To verify the
phosphorylation hypothesis, we analyzed the effect of CaMKII
antagonists, KN-62 and KN-93. The reversion of the long-term L-
Glu-inhibitory effects on nucleotide receptor response, by inhibiting
the CaMKII, confirmed the role of this protein kinase on the reported
effect. However, it is interesting to note that the immediate inhibitory
effects induced by L-Glu were not abolished by CaMKII antagonist
suggesting the involvement of other kinases. It should be noticed
that although KN-62 has been described as antagonist of human
P2X7 receptor (34) the differences in the amino acid sequence of
the first 335 residues between human and rat P2X7 receptor have
been described to change completely the sensitivity to KN-62. Thus,
in HEK 293 cells stably transfected with rat P2X7 receptor KN-62
did not antagonize neither ATP activation of cation currents nor
ethidium influx induced by ATP (35). In any case, we have
additionally verified the lack of antagonism of KN-62 on P2X7
receptor in microfluorimetric experiments where the extracellular
Ca2+ entrance induced by BzATP was not decreased in the presence
of KN-62 (data not shown).

Multiple temporarily regulated mechanisms are used to modulate
the efficiency of synaptic transmission; in the present work we
have described a new one which involves glutamate and nucleotide

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