Differential localization and regulation of stargazin-like protein, γ-8 and stargazin in the plasma membrane of hippocampal and cortical neurons

Transmembrane AMPA receptor regulatory proteins (TARPs), including stargazin/γ-2, are associated with AMPA receptors and participate in their surface delivery and anchoring at the postsynaptic membrane. TARPs may also act as a positive modulator of the AMPA receptor ion channel function; however, little is known about other TARP members except for stargazin/γ-2. We examined the synaptic localization of stargazin/γ-2 and γ-8 by immunoelectron microscopy and biochemical analysis. The analysis of sodium dodecyl sulfate-digested freeze-fracture replica labeling revealed that stargazin/γ-2 was concentrated in the postsynaptic area, whereas γ-8 was distributed both in synaptic and extra-synaptic plasma membranes of the hippocampal neuron. When a synaptic plasma membrane-enriched brain fraction was treated with Triton X-100 and separated by sucrose density gradient ultracentrifugation, a large proportion of NMDA receptor and stargazin/γ-2 was accumulated in raft-enriched fractions, whereas AMPA receptor and γ-8 were distributed in both the raft-enriched fractions and other Triton-insoluble fractions. Phosphorylation of stargazin/γ-2 and γ-8 was regulated by different sets of kinases and phosphatases in cultured cortical neurons. These results suggested that stargazin/γ-2 and γ-8 have distinct roles in postsynaptic membranes under the regulation of different intracellular signaling pathways.