Electrophysiological characterization of human keratinocytes using the patch-clamp technique

The living, not-cornified part of the epidermis consists mainly of keratino cytes. The control mechanisms of proliferation and differentiation are only partly understood. Similarly, the influence of ionic channels of the cell membrane on the proliferation of keratinocytes remains unclear. Preliminary investigations point to a relation between the opening of ionic channels a nd keratinocyte proliferation. Therefore, voltage clamp experiments were pe rformed to gain further knowledge of the electrophysiological characteristi cs of human keratinocyte cellular membranes. In-vitro cultured keratinocyte s of the cell line HaCaT were characterized by means of the voltage-clamp t echnique. As measured in the whole-cell configuration, changing the extrace llular K+ or Cl--concentration shifted the membrane potential of HaCaT cell s. Application of the patch-clamp-technique in the cell-attached and inside -out configuration revealed an ionic channel with a conductance of multiple s of 200 pS. The reversal potential of the single channel current was shift ed by substituting of intracellular Cl- by aspartate(-). Channel openings d isappeared after addition of 0.1 mM of the anion channel blocker 4',4' diis othiocyanato-stilbene-2-2' disulfonic acid (DIDS). It is concluded that thi s channel contributes to the Cl--conductance of the cellular membrane and i s a determinant of the membrane potential of human keratinocytes. This chan nel may represent a target for pharmacological manipulation of the membrane potential and possibly the growth of human keratinocytes in dermatological proliferation disorders.