Functional and morphological correlates of connexin50 expressed in Xenopuslaevis oocytes

Electrophysiological and morphological methods were used to study connexin5 0 (Cx50) expressed in Xenopus laevis oocytes. Oocytes expressing Cx50 exhib ited a new population of intramembrane particles (9.0 nm in diameter) in th e plasma membrane. The particles represented hemichannels (connexin hexamer s) because (a) their cross-sectional area could accommodate 24 +/- 3 helice s, (b) when their density reached 300-400/ mu m(2), they formed complete ch annels (dodecamers) in single oocytes, and assembled into plaques, and (c) their appearance in the plasma membrane was associated with a whole-cell cu rrent, which was activated at low external Ca2+ concentration ([Ca2+](o)), and was blocked by octanol and by intracellular acidification. The Cx50 hem ichannel density was directly proportional to the magnitude of the Cx50 Ca2 +-sensitive current. Measurements of hemichannel density and the Ca2+-sensi tive current in the same oocytes suggested that at physiological [Ca2+](o), (1-2 mM), hemichannels rarely open. In the cytoplasm, hemichannels were pr esent in similar to 0.1-mu m diameter "coated" and in larger 0.2-0.5-mu m d iameter vesicles. The smaller coated vesicles contained endogenous plasma m embrane proteins of the oocyte intermingled with 5-40 Cx50 hemichannels, an d were observed to fuse with the plasma membrane. The larger vesicles, whic h contained Cx50 hemichannels, gap junction channels, and endogenous membra ne proteins, originated from invaginations of the plasma membrane, as their lumen was labeled with the extracellular marker peroxidase. The insertion rate of hemichannels into the plasma membrane (80,000/s), suggested that an average of 4,000 small coated vesicles were inserted every second. However , insertion of hemichannels occurred at a constant plasma membrane area, in dicating that insertion by vesicle exocytosis (60-500 mu m(2) membranes/s) was balanced by plasma membrane endocytosis. These exocytotic and endocytot ic rates suggest that the entire plasma membrane of the oocyte is replaced in similar to 24 h.