The Journal of General Physiology, Vol 105, 267-287, Copyright © 1995 by The Rockefeller University Press
Irreversible inhibition of sodium current and batrachotoxin binding by a photoaffinity-derivatized local anesthetic
J McHugh, WM Mok, GK Wang and G Strichartz
Anesthesia Research Laboratories, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
We have synthesized a model local anesthetic (LA), N-(2-di-N-butyl-
aminoethyl)-4-azidobenzamide (DNB-AB), containing the photoactivatable aryl
azido moiety, which is known to form a covalent bond to adjacent molecules
when exposed to UV light (Fleet, G.W., J.R. Knowles, and R.R. Porter. 1972.
Biochemical Journal. 128:499-508. Ji, T.H. 1979. Biochimica et Biophysica
Acta. 559:39-69). We studied the effects of DNB-AB on the sodium current
(INa) under whole-cell voltage clamp in clonal mammalian GH3 cells and on
3[H]-BTX-B binding to sheep brain synaptoneurosomes. In the absence of UV
illumination, DNB-AB behaved similarly to known LAs, producing both
reversible block of peak INa (IC50 = 26 microM, 20 degrees C) and
reversible inhibition of 3[H]-BTX- B (50 nM in the presence of 0.12
microgram/liter Leiurus quinquestriatus scorpion venom) binding (IC50 = 3.3
microM, 37 degrees C), implying a noncovalent association between DNB-AB
and its receptor(s). After exposure to UV light, both block of INa and
inhibition of 3[H]-BTX-B binding were only partially reversible (INa = 42%
of control; 3[H]-BTX-B binding = 23% of control) showing evidence of a
light-dependent, covalent association between DNB-AB and its receptor(s).
In the absence of drug, UV light had less effect on INa (post exposure INa
= 96% of control) or on 3[H]-BTX-B binding (post exposure binding = 70% of
control). The irreversible block of INa was partially protected by
coincubation of DNB-AB with 1 mM bupivacaine (IC50 = 45 microM, for INa
inhibition at 20 degrees C, Wang, G.K., and S.Y. Wang. 1992. Journal of
General Physiology. 100:1003-1020), (post exposure INa = 73% of control).
The irreversible inhibition of 3[H]-BTX- B binding also was partially
protected by coincubation with bupivacaine (500 microM, 37 degrees C) (post
exposure binding = 51% of control), suggesting that the site of
irreversible inhibition of both INa and 3[H]-BTX-B binding is shared with
the clinical LA bupivacaine.
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