Confocal microscopic analysis of intracellular ph regulation in
isolated guinea pig pancreatic ducts.
Ondarza, Jos[acute]e De, and Seth R. Hootman.
Department of Physiology, Michigan State University, 108 Giltner
Hall, East Lansing, MI 48824-1101, Phone: (517) 355-6475 ext. 1242,
Fax: (517) 355-5215
APStracts 3:0060G, 1996.
pH regulation in isolated guinea pig pancreatic interlobular duct
segments loaded with the pH-sensitive fluorophore, 5-(and-6)-carboxy
-SNARF-1, acetoxymethyl ester (SNARF-1) was characterized by laser
scanning confocal microscopy. In HCO3--free medium, intracellular pH
of duct epithelial cells (pHi) fell by 0.32 +/- 0.06 pH units in the
presence of 0.5 mM amiloride and by 0.36 +/- 0.08 pH units in the
absence of Na+. In the presence of extracellular HCO3-, pHi acidified
in Na+-free, but not in amiloride-containing, medium. Superfusion
with chloride-free buffers or with buffers containing 4,4'
-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) produced a
cytosolic alkalinization of 0.13-0.22 pH units. These observations
demonstrate the presence of Na+/H+ exchange, Na+/HCO3- cotransport,
and Cl-/HCO3- exchange in guinea pig pancreatic ducts. pHi recovered
significantly from an NH4Cl pulse in HCO3--free buffers containing
amiloride and carbachol (50.4%) or amiloride and secretin (40.6%).
This recovery was blocked by the H+-ATPase inhibitor, bafilomycin A1
and by preincubation of ducts with nocodazole or cytochalasin D.
These observations suggest that a vesicular H+-ATPase augments Na+
-dependent H+ extrusion during agonist-stimulated bicarbonate
secretion, and that activation of this transport mechanism involves
cytoskeletal elements.
Received 12 June 1995; accepted in final form 28 February 1996.
APS Manuscript Number G249-5.
Article publication pending Am. J. Physiol. (Gastrointest. Liver
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 21 March 96