Little is known of the excitatory mechanisms that contribute to the tonic contraction of the corpus cavernosum smooth muscle in the flaccid state. We used patch-clamp electrophysiology to investigate a previously unidentified inward current in freshly isolated rat and human corporal myocytes. Phenylephrine (PE) contracted cells and activated whole cell currents. Outward current was identified as large-conductance Ca(2+)-activated K(+) current. The inward current elicited by PE was dependent on the Cl(-) gradient and was inhibited by niflumic acid, indicative of a Ca(2+)-activated Cl(-) (Cl(Ca)) current. Furthermore, spontaneous transient outward and inward currents (STOCs and STICs, respectively) were identified in both rat and human corporal myocytes and derived from large-conductance Ca(2+)-activated K(+) and Cl(Ca) channel activity. STICs and STOCs were inhibited by PE and A-23187, and combined 8-bromoadenosine cAMP and 8-bromoadenosine cGMP decreased their frequency. When studied in vivo, chloride channel blockers transiently increased intracavernosal pressure and prolonged nerve-evoked erections. This report reveals for the first time Cl(Ca) current in rat and human corpus cavernosum smooth muscle cells and demonstrates its key functional role in the regulation of penile erection.

Inwardly rectifying K(+) (K(IR)) currents are present in some, but not all, vascular smooth muscles. We used patch-clamp methods to examine plasticity of this current by comparing contractile and proliferative phenotypes of a clonal human vascular smooth muscle cell line. Hyperpolarization of cells under voltage clamp elicited a large inward current that was selective for K(+) and blocked by Ba(2+). Current density was greater in proliferative compared with contractile cells (-4.5 +/- 0.9 and -1.4 +/- 0.3 pA/pF, respectively; P < 0.001). RT-PCR of mRNA from proliferative cells identified transcripts for Kir2.1 and Kir2.2 but not Kir2.3 potassium channels. Western blot analysis demonstrated greater expression of Kir2.1 protein in proliferative cells, consistent with the higher current density. Proliferative cells displayed a more negative membrane potential than contractile cells (-71 +/- 2 and -35 +/- 4 mV, respectively; P< 0.001). Ba(2+) depolarized all cells, whereas small increases in extracellular K(+) concentration elicited hyperpolarization only in contractile cells. Ba(2+) inhibited [(3)H]thymidine incorporation, indicating a possible role for K(IR) channels in the regulation of proliferation. The phenotype-dependent plasticity of K(IR) channels may have relevance to vascular remodeling.

We have previously identified a human vascular smooth muscle clone that can reversibly convert between proliferative and contractile phenotypes. Here we compared receptor-channel coupling in these cells using fura-2 to monitor [Ca(2+)](i) and patch-clamp to record currents. Histamine elevated [Ca(2+)](i) in all cells and caused contraction of cells exhibiting the contractile phenotype. The rise of [Ca(2+)](i) persisted in Ca(2+)-free solution and was abolished by thapsigargin, indicating involvement of stores. Whole cell electrophysiological recording revealed that histamine evoked transient outward K(+) current, indicating functional receptor-channel coupling. The time-course and amplitude of the histamine-activated current were similar in cells of the proliferative and contractile phenotypes. Moreover, a large conductance K(+) channel was recorded in cell-attached patches and was activated by histamine as well as the Ca(2+) ionophore A-23187, identifying it as the large conductance Ca(2+)-dependent K(+) channel. This K(+) channel showed similar characteristics and activation in both proliferative and contractile phenotypes, indicating that expression was independent of phenotype. In contrast, histamine also elicited an inward Cl(-) current in some contractile cells, suggesting differential regulation of this current depending on phenotype. These studies demonstrate the usefulness of this human vascular cell clone for studying functional plasticity of smooth muscle, while avoiding complications arising from extended times in culture.

No abstract