Gap junctions in Malpighian tubules ofAedes aegypti

Abstract

SUMMARYWe present electrical, physiological and molecular evidence for substantial electrical coupling of epithelial cells in Malpighian tubules via gap junctions. Current was injected into one principal cell of the isolated Malpighian tubule and membrane voltage deflections were measured in that cell and in two neighboring principal cells. By short-circuiting the transepithelial voltage with the diuretic peptide leucokinin-VIII we largely eliminated electrical coupling of principal cells through the tubule lumen,thereby allowing coupling through gap junctions to be analyzed. The analysis of an equivalent electrical circuit of the tubule yielded an average gap-junction resistance (Rgj) of 431 kΩ between two cells. This resistance would stem from 6190 open gap-junctional channels,assuming the high single gap-junction conductance of 375 pS found in vertebrate tissues. The addition of the calcium ionophore A23187 (2 μmol l–1) to the peritubular Ringer bath containing 1.7 mmol l–1 Ca2+ did not affect the gap-junction resistance, but metabolic inhibition of the tubule with dinitrophenol (0.5 mmol l–1) increased the gap-junction resistance 66-fold,suggesting the regulation of gap junctions by ATP. Lucifer Yellow injected into a principal cell did not appear in neighboring principal cells. Thus, gap junctions allow the passage of current but not Lucifer Yellow. Using RT-PCR we found evidence for the expression of innexins 1, 2, 3 and 7 (named after their homologues in Drosophila) in Malpighian tubules. The physiological demonstration of gap junctions and the molecular evidence for innexin in Malpighian tubules of Aedes aegypti call for the double cable model of the tubule, which will improve the measurement and the interpretation of electrophysiological data collected from Malpighian tubules.