The Cl- channel CLH-3 is expressed in C. elegans oocytes, but not in surrounding sheath cells. CLH-3 is activated during oocyte meiotic maturation. CLH-3 RNAi triggers premature sheath cell ovulatory contractions suggesting that the channel functions to modulate sheath contractile activity. Oocytes are coupled to sheath cells by gap junctions. We postulated that activation of CLH-3 depolarizes the oocyte and electrically-coupled sheath cell plasma membranes and this depolarization modulates contractile activity by regulating Ca2+ influx via IP3 signaling-dependent Ca2+ channels. To test this idea, we have quantified sheath cell contractility in animals where IP3 receptor function has been disrupted by either mutation or RNAi. Sheath cells of N2 worms exhibit a basal contraction rate of 9 +/- 1 contractions/min with a displacement of 2.3 +/- 0.01 m. The force and frequency of sheath contractions increase during ovulation to a peak rate of 22 +/- 2 contractions/min with a displacement of 4.6 +/- 0.03 m. JT73 worms harbor a loss-of-function mutation in the IP3 receptor gene
itr-1, which severely inhibits sheath contractility. Rates of basal and peak ovulatory contractions were 2 +/- 1 contractions/min and 5 +/- 2 contractions/min. A similar phenotype was observed when
itr-1 expression was disrupted by RNAi. PS2368 worms harbor a gain-of-function mutation in
itr-1 in an
unc-24 genetic background. The rate of basal sheath cell contraction and the basal sheath displacement in
unc-24 worms are 8 +/- 1 contractions/min and 2.9 +/- 0.2 m. Basal sheath contractility was substantially greater in PS2368 worms. The basal rate of sheath cell contraction was 12 +/- 1 contractions/min and the contractions were more forceful exhibiting a displacement of 4.4 +/- 0.2 m. The duration of sheath contractions and the length of the sheath over which contractions were observed were also significantly higher in PS2368 worms. Mean duration of basal and ovulatory contractions were 1.0 +/- 0.1 sec and 1.5 +/- 0.1 sec in
unc-24 worms and 1.6 +/- 0.1 sec and 4.1 +/- 0.3 sec in the PS2368 strain. The length of the contracting sheath in PS2368 worms was 25.3 +/- 2.7 m and 23.1 +/- 2.0 m under basal conditions and during ovulation. In
unc-24 worms, these values were 13.4 +/- 0.9 m and 11.7 +/- 1.3m. Taken together, these results support the hypothesis that IP3 signaling plays an important role in regulating sheath cell contraction. We are currently developing methods to monitor Ca2+ levels in sheath cells and are targeting other IP3 signaling components for RNAi disruption.