Effects of the carcinogen dimethylhydrazine (DMH) on the function of rat colonic crypts

Abstract

 Rats injected with dimethylhydrazine for 5 weeks (DMH, 40 mg/kg body weight) invariably develop colonic cancer after a latency of some 10–14 weeks. Preliminary studies have suggested that Na⁺ absorption by surface colonic crypt cells is attenuated in the preneoplastic period (8–12 weeks after the first injection of DMH). The present study of glucocorticoid-treated (dexamethasone 6 mg/kg body weight, s.c. 3 days or triamcinolone 30 mg/kg body weight, s.c. 3 days) rats was undertaken to examine the ion transport properties of rat distal colon during this period in more detail. Ussing chamber studies of the distal colon and whole-cell patch-clamp measurements in surface cells, mid-crypt cells and crypt-base cells obtained from isolated crypts were performed. In Ussing chamber studies the equivalent short-circuit current inhibitable by amiloride (10 μmol/l) DMH-treated rats was about 40% of control. In addition, the hyperpolarizing effect of amiloride (10 μmol/l) on membrane voltage (V _m) was strongly attenuated in surface and mid-crypt cells of DMH-treated rats. Carbachol (CCH, 100 μmol/l), which predictably hyperpolarized surface, mid-crypt cells and crypt-base cells of control rats, had no significant effect on V _m in DMH-treated rats, but increased membrane conductance (G _m) significantly. This indicates that CCH probably activates both Cl^–and K⁺ channels in all three colonic crypt compartments in the DMH-treated rats. Forskolin (5 μmol/l), which has the most pronounced effect in crypt-base cells in control rats, depolarized V _m and enhanced G _m in all three compartments in DMH-treated rats. These data indicate that DMH profoundly alters Na⁺ and Cl^–transport in colonic crypts prior to the appearance of colonic adenocarcinoma and that these effects can be summarized as follows: (1) the Na⁺ conductance of surface cells is attenuated; (2) cells along the length of the crypt-lumen axis tend to lose their normal response to CCH and instead show simultaneous and comparable increases in K⁺and Cl^–conductances; (3) the effect of forskolin is enhanced along the entire crypt axis. As a result colonic crypt transport is shifted to predominant Cl^–secretion, findings which are characteristic of colonic carcinoma cell lines such as HT₂₉ and T₈₄ cells.