Pacemaker cells, known as interstitial cells of Cajal (ICC), generate electrical rhythmicity in the gastrointestinal tract. Pacemaker currents in ICC result from the activation of a voltage‐independent, non‐selective cation conductance, but the timing mechanism responsible for periodic activation of the pacemaker current is unknown. Previous studies suggest that pacemaking in ICC is dependent upon metabolic activity 1y1yand1 Ca2+ release from intracellular stores. We tested the hypothesis that mitochondrial Ca2+ handling may underlie the dependence of gastrointestinal pacemaking on oxidative metabolism. Pacemaker currents occurred spontaneously in cultured ICC and were associated with mitochondrial Ca2+ transients. Inhibition of the electrochemical gradient across the inner mitochondrial membrane blocked Ca2+ uptake and pacemaker currents in cultured ICC and blocked slow wave activity in intact gastrointestinal muscles from mouse, dog and guinea‐pig. Pacemaker currents and rhythmic mitochondrial Ca2+ uptake in ICC were also blocked by inhibitors of IP3‐dependent release of Ca2+ from the endoplasmic reticulum and by inhibitors of endoplasmic reticulum Ca2+ reuptake. Our data suggest that integrated Ca2+ handling by endoplasmic reticulum and mitochondria is a prerequisite of electrical pacemaking in the gastrointestinal tract.

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Considerable evidence suggests that interstitial cells of Cajal (ICC) are the pacemakers cells in gastrointestinal (GI) muscles (e.g. Langton et al. 1989; Ward et al. 1994; Huizinga et al. 1995; Thomsen et al. 1998; Koh et al. 1998; Dickens et al. 1999). Freshly isolated (Langton et al. 1998) and cultured ICC (Thomsen et al. 1998; Koh et al. 1998) generate spontaneous electrical slow waves and pacemaker currents. Absence of ICC in tissues results in loss of slow waves (Torihashi et al. 1995; and for see review Sanders, 1996). Voltage clamp studies have shown that holding cells at potentials between _80 and 0 mV does not significantly affect the frequency of spontaneous pacemaker currents, suggesting that activation of the pacemaker conductance is not voltage-dependent (Koh et al. 1998). The pacemaker currents reversed near 0 mV, were blocked by Gd 3+ , and were reduced by niflumic acid, decreased extracellular Na +

Considerable evidence suggests that interstitial cells of Cajal (ICC) are the pacemakers cells in gastrointestinal (GI) muscles (e.g.

The p53-inducible gene 3 (PIG3) is originally isolated as a p53 downstream target gene, but its function remains unknown. Here, we report a role of PIG3 in the activation of DNA damage checkpoints, after UV irradiation or radiomimetic drug neocarzinostatin (NCS). We show that depletion of endogenous PIG3 sensitizes cells to DNA damage agents, and impaired DNA repair. PIG3 depletion also allows for UV-and NCS-resistant DNA synthesis and permits cells to progress into mitosis, indicating that PIG3 knockdown can suppress intra-S phase and G2/M checkpoints. PIG3-depleted cells show reduced Chk1 and Chk2 phosphorylation after DNA damage, which may directly contribute to checkpoint bypass. PIG3 exhibited diffuse nuclear staining in the majority of untreated cells and forms discrete nuclear foci in response to DNA damage. PIG3 colocalizes with c-H2AX and 53BP1 to sites of DNA damage after DNA damage, and binds to a c-H2AX. Notably, PIG3 depletion decreases the efficient induction and maintenance of H2AX phosphorylation after DNA damage. Moreover, PIG3 contributes to the recruitment of 53BP1, Mre11, Rad50 and Nbs1 to the sites of DNA break lesions in response to DNA damage. Our combined results suggest that PIG3 is a critical component of the DNA damage response pathway and has a direct role in the transmission of the DNA damage signal from damaged DNA to the intra-S and G2/M checkpoint machinery in human cells.