ACTA HISTOCHEMICA ET CYTOCHEMICA

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One of the most important physiological roles of brain astrocytes is the maintenance of extracellular K⁺ concentration by adjusting the K⁺ influx and K⁺ efflux. The inwardly rectifying K⁺ channel Kir4.1 has been identified as an important member of K⁺ channels and is highly concentrated in glial endfeet membranes. Aquaporin (AQP) 4 is another abundantly expressed molecule in astrocyte endfeet membranes. We examined the ultrastructural localization of Kir4.1, AQP4, α1-syntrophin, and β-spectrin molecules to understand the functional role(s) of Kir4.1 and AQP4. Immunogold electron microscopy of these molecules showed that the signals of these molecules were present along the plasma membranes of astrocyte endfeet. Double immunogold electron microscopy showed frequent co-localization in the combination of molecules of Kir4.1 and AQP4, Kir4.1 and α1-syntrophin, and AQP4 and α1-syntrophin, but not those of AQP4 and β-spectrin. Our results support biochemical evidence that both Kir4.1 and AQP4 are associated with α1-syntrophin by way of postsynaptic density-95, Drosophila disc large protein, and the Zona occludens protein I protein-interaction domain. Co-localization of AQP4 and Kir4.1 may indicate that water flux mediated by AQP4 is associated with K⁺ siphoning.

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We examined the expression and intracellular localization of vesicle-associated membrane protein 2 (VAMP2) during the differentiation of skeletal muscle cells by immunofluorescence microscopy. In isolated single myofibers, VAMP2 was expressed in quiescent satellite cells, downregulated in proliferating myoblastic cells, and re-expressed with differentiation. In the myoblastic cell line C2C12, VAMP2 was expressed at a low level in the proliferating stage, and then increased after differentiation into myotubes. Based on these results, we propose that VAMP2 can be used as a molecular marker for both quiescent satellite cells and myotubes, but not for proliferating myoblasts. We also found the partial colocalization of VAMP2 with transferrin- or Rab11-labeled vesicles in myotubes, suggesting a role of VAMP2 in the trafficking of recycling endosomes.

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