Expression of the cystic fibrosis transmembrane conductance regulator in rat spermatids: Implication for the site of action of antispermatogenic agents

Abstract

To establish whether cystic fibrosis transmembrane conductance regulator (CFTR) is functionally expressed in the testis, we subjected spermatogenic cells from rat testes to analysis of CFTR mRNA, protein and channel activity. CFTR mRNA was detected in the testes of mature but not immature rats using reverse transcription-polymerase chain reaction analysis. Western blot analysis performed with a CFTR specific antibody revealed immunoreactivity in the membrane extract of spermatogenic cells. Immunohistochemical studies localized CFTR in round and elongated spermatids, but not in the fully developed spermatozoa. Using a whole-cell patch clamp technique, we recorded an inward current activated by intracellular cAMP (100 μmol/l) in round spermatids. The current displayed a linear I/V relationship and was inhibited by diphenylamine-2-carboxylate (DPC), a chloride channel blocker. Transfection of the rat germ cell CFTR cDNA into human embryonic kidney (HEK) 293 cells caused the expression of a cAMP-activated chloride current with CFTR characteristics. The current was completely blocked by the antispermatogenic agents 1-(2,4-dichlorobenzyl)-indazole-3-carboxylic acid, lonidamine (500 υmol/l) and 1-(2,4-dichlorobenzyl)-indazole-3-acrylic acid, AF2785 (250 υmol/l). These results taken together provide evidence that CFTR is differentially expressed in spermatids during spermiogenesis. We speculate that CFTR may interact with aquaporin to bring about cytoplasmic volume contraction which is an essential feature of spermiogenesis.