Summary The Na
+ requirement for active, electrogenic Cl
– absorption by
Amphiuma small intestine was studied by tracer techniques and double-barreled Cl
–-sensitive microelectrodes. Addition of Cl
– to a Cl
–-free medium bathing
in vitro intestinal segments produced a saturable (
K
m
=5.4
mm) increase in shortcircuit current (
I
sc) which was inhibitable by 1
mm SITS. The selectivity sequence for the anion-evoked current was Cl
–=Br
–>SCN
–>NO
3
–
>F
–=I
–. Current evoked by Cl
– reached a maximum with increasing medium Na concentration (
K
m
=12.4
mm). Addition of Na
+, as Na gluconate (10
mm), to mucosal and serosal Na
+-free media stimulated the Cl
– current and simultaneously increased the absorptive Cl
– flux (
J
ms
Cl
) and net flux (
J
net
Cl
) without changing the secretory Cl
– flux (
J
sm
Cl
). Addition of Na
+ only to the serosal fluid stimulated
J
ms
Cl
much more than Na
+ addition only to the mucosal fluid in paired tissues. Serosal DIDS (1
mm) blocked the stimulation. Serosal 10
mm Tris gluconate or choline gluconate failed to stimulate
J
ms
Cl
. Intracellular Cl
– activity (
a
Cl
i
) in villus epithelial cells was above electrochemical equilibrium indicating active Cl
– uptake. Ouabain (1
mm) eliminated Cl
– accumulation and reduced the mucosal membrane potential
m
over 2 to 3 hr. In contrast, SITS had no effect on Cl
– accumulation and hyperpolarized the mucosal membrane. Replacement of serosal Na
+ with choline eliminated Cl
– accumulation while replacement of mucosal Na
+ had no effect. In conclusion by two independent methods active electrogenic Cl
– absorption depends on serosal rather than mucosal Na
+. It is concluded that Cl
– enters the cell via a primary (rheogenic) transport mechanism. At the serosal membrane the Na
+ gradient most likely energizes H
+ export and regulates mucosal Cl
– accumulation perhaps by influencing cell pH or HCO
3
–
concentration.
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