Anoctamin 6 Contributes to Cl- Secretion in Accessory Cholera Enterotoxin (Ace)-stimulated Diarrhea: AN ESSENTIAL ROLE FOR PHOSPHATIDYLINOSITOL 4,5-BISPHOSPHATE (PIP2) SIGNALING IN CHOLERA.

Accessory cholera enterotoxin (Ace) of Vibrio cholerae has been shown to contribute to diarrhea. However, the signaling mechanism and specific type of Cl- channel activated by Ace are still unknown. We have shown here that the recombinant Ace protein induced ICl of apical plasma membrane, which was inhibited by classical CaCC blockers. Surprisingly, an Ace-elicited rise of current was neither affected by ANO1 (TMEM16A)-specific inhibitor T16A(inh)-AO1(TAO1) nor by the cystic fibrosis transmembrane conductance regulator (CFTR) blocker, CFTR inh-172. Ace stimulated whole-cell current in Caco-2 cells. However, the apical ICl was attenuated by knockdown of ANO6 (TMEM16F). This impaired phenotype was restored by re-expression of ANO6 in Caco-2 cells. Whole-cell patch clamp recordings of ANO currents in HEK293 cells transiently expressing mouse ANO1-mCherry or ANO6-GFP confirmed that Ace induced Cl- secretion. Application of Ace produced ANO6 but not the ANO1 currents. Ace was not able to induce a [Ca2+]i rise in Caco-2 cells, but cellular abundance of phosphatidylinositol 4,5-bisphosphate (PIP2) increased. Identification of the PIP2-binding motif at the N-terminal sequence among human and mouse ANO6 variants along with binding of PIP2 directly to ANO6 in HEK293 cells indicate likely PIP2 regulation of ANO6. The biophysical and pharmacological properties of Ace stimulated Cl- current along with intestinal fluid accumulation, and binding of PIP2 to the proximal KR motif of channel proteins, whose mutagenesis correlates with altered binding of PIP2, is comparable with ANO6 stimulation. We conclude that ANO6 is predominantly expressed in intestinal epithelia, where it contributes secretory diarrhea by Ace stimulation in a calcium-independent mechanism of RhoA-ROCK-PIP2 signaling.

[Lipid peroxidation and the antioxidant blood system in dynamics of pestilential and choleraic intoxication]. / Lipidperoksidatsiia i antioksidantnaia sistema krovi v dinamike chumnoi i kholernoi intoksikatsii.

Lipid peroxidation (LPO) and activity of antioxidant system were studied spectrophotometrically in white rats developing pestilential and choleraic intoxication achieved by intraperitoneal injection of plague autolysate of vaccine EB strain in dose equivalent to DL50, cholerain endotoxin in doses DL50 and DL25 as well as combined effect of choleraic endo- and enterotoxins. With progression of the intoxication, the levels of diene conjugates and malonic dialdehyde in blood plasma and erythrocytes rose. LPO activation in plague intoxication arose in high activity of SOD and blood catalase. In choleraic intoxication the activity of the above enzymes progressively lowered.

On the role of intramural nerves in the pathogenesis of cholera toxin-induced intestinal secretion.

Intestinal secretion was produced in anesthetized cats and rats by exposing isolated intestinal segments to cholera enterotoxin. Giving, for example, tetrodotoxin, a nerve-conduction-blocking agent, or adding lidocaine, a local anesthetic agent, to the solution in the intestinal segments markedly inhibited the rate of choleraic secretion, and in most experiments a net absorption of fluid was observed. The results suggest that intramural nervous mechanisms are involved in the pathogenesis of choleraic secretion.

Berberine in toxin-induced experimental cholera.

1. Oral administration of berberine to infant rabbits 18-24 h before the intraintestinal administration of choleragenic toxins, arrests diarrhoea or significantly prolongs the survival time.2. The use of berberine in the treatment of clinical cholera is further justified.3. Berberine is an antidiarrhoeal drug and the host tissues play a major part in the control of diarrhoeal symptoms.