Hypercontractility and impaired sildenafil relaxations in the BKCa channel deletion model of erectile dysfunction

Abstract

Erectile dysfunction (ED) can be elicited by a variety of pathogenic factors, particularly impaired formation of and responsiveness to nitric oxide (NO) and the downstream effectors soluble guanylate cyclase (sGC) and cGMP-dependent protein kinase I (PKGI). One important target of PKGI in smooth muscle is the large-conductance, Ca2+ -activated potassium (BKCa) channel. In our previous report (42), we demonstrated that deletion of the BKCa channel in mice induced force oscillations and led to reduced nerve-evoked relaxations and ED. In the current study, we used this ED model to explore the role of the BKCa channel in the NO/sGC/PKGI pathway. Electrical field stimulation (EFS)-induced contractions of corpus cavernosum smooth muscle strips were significantly enhanced in the absence of BKCa channel function. In strips precontracted with phenylephrine, EFS-induced relaxations were converted to contractions by inhibition of sGC, and this was further enhanced by loss of BK channel function. Sildenafil-induced relaxations were decreased to a similar extent by inhibition of sGC or BKCa channels. At concentrations >1 microM, sildenafil caused relaxations independent of inhibition of sGC or BKCa channels. Sildenafil did not affect the enhanced force oscillations that were induced by the loss of BKCa channel function. Yet, these oscillations could be completely eliminated by blocking L-type voltage-dependent Ca2+ channels (VDCCs). These results suggest that therapeutically relevant concentrations of sildenafil act through cGMP and BKCa channels, and loss of BKCa channel function leads to hypercontractility, which depends on VDCCs and cannot be modified by the cGMP pathway.

Fig. 1.

Electrical field stimulation (EFS)-induced contractions are increased in corpus cavernosum smooth muscle (CCSM) strips from Slo^−/− mice. A: representative recordings from a Slo^+/+ (left) and a Slo^−/− (right) CCSM strip of EFS-induced contractions with increasing frequencies (1, 2, 3.5, 5, 7.5, 10, 12.5, 15, 20, 30, 40, and 50 Hz) every 3 min. B: representative recordings from Slo^+/+ (left) and Slo^−/− (right) CCSM strips of EFS-induced contractions with continuous 30-Hz stimulation every minute before and after adding iberiotoxin (IBTX). C: relationship between stimulation frequency and force in CCSM strips from Slo^+/+ and a Slo^−/− mice. D: effect of IBTX on continuous 30-Hz EFS-induced contractions, normalized to values before IBTX; n, no. of CCSM strips; N, no. of mice. *P < 0.05 vs. Slo^+/+; ns, Not significant.

Fig. 2.

Inhibition of soluble guanylate cyclase (sGC) turns EFS-induced relaxations into contractions. A: sample recordings from a phenylephrine (PE)-precontracted Slo^+/+ (left) and a Slo^−/− (right) CCSM strip and EFS-induced relaxations lasting 2 and 60 s. B: same as in A but after adding ODQ to the bath. C and D: summary of relaxations and contractions induced by the 2-s (C) and 60-s (D) EFS shown in A and B; n, no. of CCSM strips from 3 mice; *P < 0.05 vs. Slo^+/+.

Fig. 3.

Sildenafil-mediated suppression of PE-induced contractions is diminished in Slo^−/−. A and B: representative PE-induced contractions recorded from a Slo^+/+ (left) and a Slo^−/− (right) CCSM strip before (A) and after (B) adding sildenafil (SIL). C: summary of the SIL-mediated reduction of PE-induced contractions as shown in A and B; n, no. of CCSM strips; N, no. of mice; P < 0.01 vs. untreated (*) and vs. Slo^+/+ + SIL (#).

Fig. 4.

Sildenafil-induced relaxation of PE-precontracted CCSM strips is reduced in Slo^−/−. A and B: sample recordings from a PE-precontracted Slo^+/+ (left) and a Slo^−/− (right) CCSM strip in the absence (A) and presence (B) of IBTX that represent the SIL-induced relaxation. C and D: average oscillation amplitudes and frequency, respectively, of Slo^+/+ and a Slo^−/− CCSM strip in the absence and presence of IBTX, before and after adding SIL. E, average %SIL-induced relaxation of PE-precontracted strips shown in A and B. F: dose-response curves of Slo^+/+ and Slo^−/− CCSM strips to SIL from 1 nM to 10 μM in the absence and presence of ODQ; n, no. of CCSM strips; N, no. of mice. P < 0.05, Slo^+/+ vs. all others (*), untreated vs. sildenafil treated (#), Slo^−/− vs. Slo^−/− + ODQ (£), and Slo^+/+ + ODQ vs. Slo^−/− + ODQ ($).

Fig. 5.

Blocking of voltage-dependent Ca²⁺ channels (VDCC) inhibits force oscillations in Slo^−/− CCSM strips and in Slo^+/+ with IBTX. A and B: representative recordings from a PE-precontracted Slo^+/+ (left) and a Slo^−/− (right) CCSM strip in the absence (A) and presence (B) of IBTX that demonstrate the effect of blocking VDCC with nisoldipine. C: average oscillation amplitudes of Slo^+/+ and a Slo^−/− CCSM strip in the absence and presence of IBTX, before and after adding nisoldipine. D: summary of total relaxation of PE-precontracted CCSM strips induced by nisoldipine as shown in A and B. NIS, nisoldipine; N, no. of mice; *P < 0.05 vs. Slo^+/+ untreated.