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Re‐evaluating the Na + conductance of adult rat alveolar type II pneumocytes: evidence for the involvement of cGMP‐activated cation channels
Author(s) -
Kemp Paul J.,
Kim KwangJin,
Borok Zea,
Crandall Edward D.
Publication year - 2001
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.2001.t01-1-00693.x
Subject(s) - amiloride , chemistry , patch clamp , biophysics , conductance , sodium , endocrinology , biochemistry , biology , receptor , mathematics , organic chemistry , combinatorics
1 Alveolar epithelial type II pneumocytes were isolated and purified from adult rat lung by elastase digestion and differential adhesion, and cultured in serum‐free medium for ∼2 days on glass coverslips for subsequent patch‐clamp studies employing symmetrical sodium isethionate solutions. 2 Whole‐cell Na + currents exhibited essentially linear current‐voltage relationships which were mildly inhibited (by ∼25 %) by 10 μ m amiloride. In contrast, 1 m m Zn 2+ inhibited the currents by ∼55 % with an IC 50 of ∼134 μ m and maximal blockade achieved between 5 and 10 m m . The effects of Zn 2+ and amiloride were additive, and independent of the order of blocker addition. 3 Gd 2+ , Zn 2+ and La 3+ at 10 m m were all effective at rapidly, reversibly and significantly blocking the amiloride‐insensitive currents by ∼60 %. In contrast, Ni 2+ was a very weak inhibitor (30 % inhibition at 10 m m ). 4 Pimozide (10 μ m ) caused inhibition of whole‐cell cation conductance by ∼55 %. The inhibitory effect of pimozide was concentration dependent with an IC 50 of ∼1 μ m and was maximally effective between 10 and 30 μ m . Sequential addition of Zn 2+ and pimozide, in either order, revealed no overlapping inhibitory effect on the amiloride‐insensitive conductance, and supported the notion that the Zn 2+ ‐ and pimozide‐sensitive currents are identical. 5 The amiloride‐insensitive, Zn 2+ ‐blockable conductance was characterised by a Na + /K + permeability ratio ( P Na / P K ) of 0.73 ± 0.02. 6 8Br‐cGMP (100 μ m ), a membrane‐permeable analogue of cGMP, evoked a robust activation of whole‐cell cation conductance to 220 % of control. This activation was apparent in either the absence or the presence of 10 μ m amiloride, but was completely abolished in the presence of Zn 2+ . 7 These data support the in vivo and in situ observations of a substantial amiloride‐resistant Na + conductance, demonstrate directly that cyclic nucleotide‐gated non‐selective cation channels are functionally expressed in alveolar epithelial type II cells, and suggest that these channels may contribute to the fluid‐reabsorptive driving force in adult lung.