THE EFFECT OF SECRETAGOGUES ON ION CONDUCTANCES OF IN-VITRO PERFUSED, ISOLATED RABBIT COLONIC CRYPTS

Several secretagogues were used in this study, including those which enhance intracellular cyclic adenosine monophosphate (cAMP) production, as well as others which elevate intracellular Ca2+ activity and are known to increase Cl- secretion in the intact colon and in colonic carcinoma cell lines. They were examined with respect to their effects on electrophysiological properties in isolated rabbit distal colonic crypts. Crypts were dissected manually and perfused in vitro. Transepithelial voltage (V-te), transepithelial resistance (R(te)), membrane voltage across the basolateral membrane (V-bl), and fractional basolateral membrane resistance (FR(bl)), were estimated. Basolateral prostaglandin E(2) (PGE(2), greater than or equal to 0.1 mu mol/l), vasoactive intestinal peptide (VIP, 1 nmol/l) and adenosine (0.1 mmol/l) induced an initial depolarisation and a secondary partial repolarisation of V-bl In the case of adenosine, the initial depolarization of V-bl was by 31 +/- 2 mV (n = 47). R(te) fell significantly from 16.4 +/- 3.6 to 14.2 +/- 3.7 Ohm . cm(2) (n = 6), and FR(bl) increased significantly from 0.11 +/- 0.02 to 0.51 +/- 0.10 (n = 6). In the second phase the repolarisation of V-bl amounted 11 +/- 2 mV (n = 47) and a steady-state (V-bl) of -51 +/- 2 mV (n = 47) was reached. R(te) fell further and significantly to a steady-state value of 12.4 +/- 3.8 Ohm . cm(2) (n = 6) and FR(bl) fell significantly to 0.42 +/- 0.13 (n = 6). In 30% of the experiments, a transient hyperpolarisation of V-bl by 8 +/- 2 mV (n = 14) was seen during wash out of adenosine. In the presence of adenosine, but not under control conditions, lowering of luminal Cl- concentration from 120 to 32 mmol/l depolarised V-bl significantly by 8 +/- 1 mV (n = 9). Basolateral ATP and ADP (0.1 mmol/l) led to a short initial depolarisation followed by a sustained and significant hyperpolarisation by 6 +/- 2 mV (n = 27) and 5 +/- 4 mV (n = 8), respectively. Carbachol (CCH) hyperpolarised V-bl in a concentration-dependent manner. At 100 mu mol/l (bath) the hyperpolarisation was by 14 +/- 2 mV (n = 11) and FR(bl) fell slightly. Neurotensin (greater than or equal to 10 nmol/l), isoproterenol (greater than or equal to 10 mu mol/l) and uridine 5'-triphosphate (UTP, 0.1 mmol/l) had no effect. It is concluded that PGE(2), VIP and adenosine upregulate sequentially a luminal Cl- conductance and a basolateral K+ conductance by increasing intracellular cAMP concentration. Ca2+ mobilising hormones such as ATP, ADP, and CCH increase the basolateral K+ conductance, while the effect on luminal Cl- conductance appears to be very limited.