Significance: The peristaltic reflex elicits colonic migrating motor complexes (CMMCs) that are thought to be generated by enteric excitatory neurons stimulating smooth muscle cells (SMCs). We found that atropine did not block CMMCs and present a concept showing that poststimulus excitatory responses following nitrergic responses in interstitial cells of Cajal (ICC) are responsible for initiation of CMMCs. Ca ²⁺ transients in ICC are inhibited by nitrergic stimulation. After the inhibitory period, Ca ²⁺ transients are increased, activating currents that are conducted to SMCs and initiate propagated contractions. Poststimulus excitation generated by ICC explains the properties of CMMCs. This is important because current therapeutic manipulation of colonic transit is typically directed at strengthening cholinergic responses, whereas targeting nitrergic responses may be more suitable.

Ingested food is received, mixed, and ground into chyme by distinct gastric motility patterns. Diabetes impairs gastric muscle function, but the mechanisms underlying diabetes-induced gastric muscle dysfunction are unknown. Here, we compared the expression and phosphorylation of Ca²⁺sensitization and contractile proteins in human gastric muscles from obese nondiabetic and diabetic patients. We also compared the spontaneous phasic contractions and the contractile responses evoked by electrical field stimulation of cholinergic motor neurons. Fundus and antrum muscles were obtained from sleeve gastrectomies and were used in in vitro myobath contractile studies and for capillary electrophoresis and immunodetection ofγ-actin, CPI-17, pT38-CPI-17, MYPT1, pT853-MYPT1, pT696-MYPT1, myosin light chain (MYL9), pS19-MYL9, myosin light chain kinase (MYLK), protein phosphatase-1δ(PP1δ), and Rho-associated kinase (ROCK2). In diabetic fundus muscles, MYLK, ROCK2, and PP1δexpression was unchanged; MYPT1 and CPI-17 expression was decreased; and the pT853/MYPT1 and pT38/CPI-17 ratios, but not the pT696/MYPT1 ratio, were increased. Although MYL9 expression was increased, the pS19/MYL9 ratio was unchanged in diabetic fundus muscles. In diabetic antrum muscles, MYLK and MYL9 expression was unchanged, but ROCK2, CPI-17, and PP1δexpression was decreased. The pT38/CPI-17 ratio was unchanged, while the pS19/MYL9, pT853/MYPT1, and pT696/MYPT1 ratios were decreased, consistent with the reduced ROCK2 expression. The frequencies of spontaneous phasic contractions from nondiabetic and diabetic gastric fundus and antrum muscles did not significantly differ from each other, regardless of age, sex, or diabetic status. The fold increases in the contractions of diabetic fundus and antrum muscles in response to increased frequencies of electrical field stimulation were significantly lower compared to nondiabetic fundus and antrum muscles. The altered contractile responses and the protein expression and phosphorylation in gastric muscles of obese patients with diabetes illustrate the importance of understanding how smooth muscle Ca²⁺sensitization mechanisms contribute to gastric motility.

Adenosine 5′-triphosphate (ATP) is released in the bladder lumen during filling. Urothelial ATP is presumed to regulate bladder excitability. Urinary ATP is suggested as a urinary biomarker of bladder dysfunctions since ATP is increased in the urine of patients with overactive bladder, interstitial cystitis or bladder pain syndrome. Altered urinary ATP might also be associated with voiding dysfunctions linked to disease states associated with metabolic syndrome. Extracellular ATP levels are determined by ATP release and ATP hydrolysis by membrane-bound and soluble nucleotidases (s-NTDs). It is currently unknown whether s-NTDs regulate urinary ATP. Using etheno-ATP substrate and HPLC-FLD detection techniques, we found that s-NTDs are released in the lumen of ex vivo mouse detrusor-free bladders. Capillary immunoelectrophoresis by ProteinSimple Wes determined that intraluminal solutions (ILS) collected at the end of filling contain ENTPD3 > ENPP1 > ENPP3 ≥ ENTPD2 = NT5E = ALPL/TNAP. Activation of adenylyl cyclase with forskolin increased luminal s-NTDs release whereas the AC inhibitor SQ22536 had no effect. In contrast, forskolin reduced and SQ22536 increased s-NTDs release in the lamina propria. Adenosine enhanced s-NTDs release and accelerated ATP hydrolysis in ILS and lamina propria. Therefore, there is a regulated release of s-NTDs in the bladder lumen during filling. Aberrant release or functions of urothelial s-NTDs might cause elevated urinary ATP in conditions with abnormal bladder excitability.

Elevations in the intracellular Ca²⁺concentration activate the serine/threonine protein kinase Ca²⁺/calmodulin-dependent protein kinase II (CaM kinase II). We tested the hypothesis that increased sarco(endo)plasmic reticulum Ca²⁺-ATPase activity by phospholamban (PLB) phosphorylation contributes to smooth muscle relaxation by elevating the sarcoplasmic reticulum (SR) Ca²⁺load and increasing the frequency of Ca²⁺release events from the SR. We have previously shown that caffeine or sodium nitroprusside (SNP) relaxes murine gastric fundus smooth muscles and increases PLB phosphorylation by CaM kinase II. These findings suggest that an increased SR Ca²⁺load increases the frequency of Ca²⁺transients from the SR and results in PLB phosphorylation by CaM kinase II, contributing to caffeine- or SNP-induced relaxation. The aim of the present study was to investigate the effects of SNP on CaM kinase II and PLB phosphorylation in gastric antrum smooth muscles. SNP or 8-bromo-cGMP decreased the basal tone and amplitudes of spontaneous phasic contractions and activated CaM kinase II. SNP-induced relaxation and CaM kinase II activation were blocked by [1,2,4]oxadizolo-[4,3α]quinoxaline-1-one (ODQ) and inhibited by cyclopiazonic acid (CPA) or KN-93. SNP also increased PLBSer¹⁶and PLBThr¹⁷phosphorylation. Both PLBSer¹⁶and Thr¹⁷phosphorylation were ODQ sensitive. However, only PLBThr¹⁷phosphorylation was inhibited by CPA or KN-93. These results suggest that CaM kinase II activation and PLB phosphorylation participate in the relaxant effect of SNP on murine gastric antrum smooth muscles through a nitric oxide/guanylyl cyclase/cGMP pathway.