Intracellular pH regulation in colonocytes of rat proximal colon.
The regulation of intracellular pH (pH(i)) in colonocytes of the rat proximal colon has been investigated using the pH-sensitive dye BCECF and compared with the regulation of pH(i) in the colonocytes of the distal colon. The proximal colonocytes in a HEPES-buffered solution had pH(i)=7.24+/-0.04 and removal of extracellular Na(+) lowered pH(i) by 0.24 pH units. Acid-loaded colonocytes by an NH(3)/NH(4)(+) prepulse exhibited a spontaneous recovery that was partially Na(+)-dependent and could be inhibited by ethylisopropylamiloride (EIPA). The Na(+)-dependent recovery rate was enhanced by increasing the extracellular Na(+) concentration and was further stimulated by aldosterone. In an Na(+)- and K(+)-free HEPES-buffered solution, the recovery rate from the acid load was significantly stimulated by addition of K(+) and this K(+)-dependent recovery was partially blocked by ouabain. The intrinsic buffer capacity of proximal colonocytes at physiological pH(i) exhibited a nearly 2-fold higher value than in distal colonocytes. Butyrate induced immediate colonocyte acidification that was smaller in proximal than in distal colonocytes. This acidification was followed by a recovery phase that was both EIPA-sensitive and -insensitive and was similar in both groups of colonocytes. In a HCO(3)(-)/CO(2)-containing solution, pH(i) of the proximal colonocytes was 7.20+/-0.04. Removal of external Cl(-) caused alkalinization that was inhibited by DIDS. The recovery from an alkaline load induced by removal of HCO(3)(-)/CO(2) from the medium was Cl(-)-dependent, Na(+)-independent and blocked by DIDS. Recovery from an acid load in EIPA-containing Na(+)-free HCO(3)(-)/CO(2)-containing solution was accelerated by addition of Na(+). Removal of Cl(-) inhibited the effect of Na(+). In summary, the freshly isolated proximal colonocytes of rats express Na(+)/H(+) exchanger, H(+)/K(+) exchanger ((H(+)-K(+))-ATPase) and Na(+)-dependent Cl(-)/HCO(3)(-) exchanger that contribute to acid extrusion and Na(+)-independent Cl(-)/HCO(3)(-) exchanger contributing to alkali extrusion. All of these are likely involved in the regulation of pH(i) in vivo. Proximal colonocytes are able to maintain a more stable pH(i) than distal cells, which seems to be facilitated by their higher intrinsic buffer capacity.
Vanecková
I
I
Institute of Physiology, Czech Academy of Sciences, Prague, Czech Republic.
Vylitová-Pletichová
M
M
Beskid
S
S
Zicha
J
J
Pácha
J
J
D000643:Ammonium Chloride; D000818:Animals; D017920:Antiporters / Q000737:chemistry; D001639:Bicarbonates / Q000737:chemistry*; D002021:Buffers; D002087:Butyrates; D002478:Cells, Cultured; D003106:Colon / Q000737:chemistry*; D017506:H(+)-K(+)-Exchanging ATPase / Q000737:chemistry; D006531:HEPES; D006863:Hydrogen-Ion Concentration; D008297:Male; D018408:Patch-Clamp Techniques; D011522:Protons; D051381:Rats; D017208:Rats, Wistar; D012965:Sodium Chloride
D017920:Antiporters; D001639:Bicarbonates; D002021:Buffers; D002087:Butyrates; D011522:Protons; D000643:Ammonium Chloride; D012965:Sodium Chloride; D017506:H(+)-K(+)-Exchanging ATPase; D006531:HEPES