Disodium cromoglycate reverses colonic visceral hypersensitivity and influences colonic ion transport in a stress-sensitive rat strain

Abstract

The interface between psychiatry and stress-related gastrointestinal disorders (GI), such as irritable bowel syndrome (IBS), is well established, with anxiety and depression the most frequently occurring comorbid conditions. Moreover, stress-sensitive Wistar Kyoto (WKY) rats, which display anxiety- and depressive-like behaviors, exhibit GI disturbances akin to those observed in stress-related GI disorders. Additionally, there is mounting preclinical and clinical evidence implicating mast cells as significant contributors to the development of abdominal visceral pain in IBS. In this study we examined the effects of the rat connective tissue mast cell (CTMC) stabiliser, disodium cromoglycate (DSCG) on visceral hypersensitivity and colonic ion transport, and examined both colonic and peritoneal mast cells from stress-sensitive WKY rats. DSCG significantly decreased abdominal pain behaviors induced by colorectal distension in WKY animals independent of a reduction in colonic rat mast cell mediator release. We further demonstrated that mast cell-stimulated colonic ion transport was sensitive to inhibition by the mast cell stabiliser DSCG, an effect only observed in stress-sensitive rats. Moreover, CTMC-like mast cells were significantly increased in the colonic submucosa of WKY animals, and we observed a significant increase in the proportion of intermediate, or immature, peritoneal mast cells relative to control animals. Collectively our data further support a role for mast cells in the pathogenesis of stress-related GI disorders.

Figure 1. Disodium cromoglycate inhibits visceral hypersensitivity in stress-prone Wistar Kyoto rats.

Colorectal distension-induced pain behaviors in Wistar Kyoto (WKY) animals were significantly greater than those observed in Sprague Dawley (SD) animals (A, threshold; B, total pain behaviours). Pre-treatment with 50 mg kg^-1 disodium cromoglycate (DSCG) significantly increased the pressure required to induce the first pain behaviour (C, threshold) and decreased the total number of pain behaviours (D) in WKY animals. n= 7-8. * p<0.05, ** p <0.01, versus SD. # p <0.05, ## p <0.01, versus vehicle.

Figure 2. Stimulated release of colonic mast cell mediators was insensitive to inhibition by disodium cromoglycate.

Compound 48/80 (10 µg ml^-1)-stimulated RMCPII release was significantly greater from Wistar Kyoto (WKY) colons relative to Sprague Dawley (SD; A) tissues and was insensitive to inhibition by DSCG (C). Similarly, histamine release was neither elevated from WKY colon relative to SD tissues (B), nor sensitive to inhibition by DSCG (D). n= 7-8. ** p <0.01, versus SD.

Figure 3. Wistar Kyoto rat colon displays altered sensitivity to compound 48/80 and disodium cromoglycate in vitro.

The change in short-circuit current (Isc) induced by compound 48/80 was significantly decreased in Wistar Kyoto (WKY) colon, and sensitive to inhibition by disodium cromoglycate (DSCG; A). Though WKY and Sprague Dawley tissues responded in a similar manner to forskolin stimulation, only the WKY response was sensitive to inhibition by DSCG. n=7-8, ** p <0.01, *** p<0.001, versus SD. # p <0.05, ## p <0.01, versus vehicle.

Figure 4. Changes in submucosal mast cell number and peritoneal mast cell classification in Wistar Kyoto rats.

Colons from Wistar Kyoto (WKY) animals displayed a significantly greater number of Alcian blue (A and B, black arrow head) and RMCPII (C and D, white arrow head) positive submucosal connective tissue mast cells relative to Sprague Dawley (SD; insert) animals (* p<0.05, ** p<0.01 versus SD, n=10-15). Purified peritoneal mast cells isolated from SD animals were predominantly safranin positive (F, black arrow head), and therefore considered CTMC, while equal proportions of those isolated from WKY animals were either safranin positive or Alcian blue/safranin positive (F; white arrow heads), representing CTMC and intermediate mast cells respectively. n=4. * p<0.05, versus s⁺. # p<0.05, ## p<0.01, versus SD.