Differential expression of PKC isoforms in fresh and cultured rabbit CCD

Growth of rabbit cortical collecting duct (CCD) cells in primary culture results in a phenotype in which there is a stable stimulation of Na+ transport by arginine vasopressin. Our objective was to determine whether this altered phenotype was associated with altered expression of protein kinase C (PKC) isoforms. Western blot analysis of extracted proteins and reverse transcription-polymerase chain reaction analysis of extracted RNA showed expression of PKC-alpha, -epsilon, and -zeta in microdissected CCD segments and in fresh and cultured immunodissected CCD cells. These techniques also suggested that the rabbit CCD expresses PKC-eta- and -theta-like isoforms, which have not been identified in this species. Growth of CCD cells in primary culture produced no apparent change in the level of expression of PKC-alpha, -epsilon, or -zeta; however, the theta-like isoform was strongly expressed in fresh CCDs but only weakly expressed in cultured CCDs. The putative eta-isoform was more heavily expressed in cultured than in fresh CCD cells. The consequences of altered expression of these two isoforms in fresh and cultured rabbit CCD remain unknown, but the possibility exists that they are involved in the altered arginine vasopressin response of cultured cells.