CELL-ATTACHED PATCH CLAMPING OF THE INTACT RABBIT CILIARY EPITHELIUM

Following thorough removal of adhering aqueous humor, we have succeede d in patch clamping the intact rabbit ciliary epithelium in the cell-a ttached and inside-out excised-patch modes. Rapidly fluctuating curren ts ('chatter activity') were observed during recordings conducted for as long as 1 h. Chatter activity did not reflect seal instability sinc e interconversion was noted between chatter activity and transitions b etween stable open and closed states, excision of patches into the bat h was associated with substantial shifts in the reversal potential, an d chatter activity could be triggered by sustained hyperpolarization, but was insensitive to stretch. The chatter channel was identified as cation-nonselective from the reversal potentials both in the cell-atta ched and excised-patch modes. The channel's kinetics were similar to t hose of the cGMP-activated phototransduction channel. The results of P CR amplifications of fragments in cDNA libraries from both human cilia ry body and human nonpigmented ciliary epithelial (NPE) cells indicate d that human ciliary epithelial cells transcribe message for the retin al phototransduction channel. The possible role of the phototransducti on channel in expressing chatter activity was further explored by perf using preparations with a known activator of that channel (cGMP) and w ith a known inhibitor (L-cis-diltiazem). Neither agent significantly a ffected chatter behavior. We conclude that: (1) this is the first demo nstration of the feasibility of patch-clamping the intact ciliary epit helium; (2) the NPE cells display chatter activity arising from rapidl y fluctuating transitions of a cation-nonselective channel; (3) NPE ce lls can transcribe message for the cation-nonselective phototransducti on channel; and (4) if the observed chatter activity is from a homolog ue of the phototransduction channel, the homologue is pharmacologicall y distinct.