Conductance and kinetic properties of single nicotinic acetylcholine receptor channels in rat sympathetic neurones.
1. The unitary conductance of nicotinic acetylcholine (ACh) receptor channels in rat sympathetic neurones has been studied. Conductance estimates varied from 26-48 pS with a mean of 36.8 pS in 1 mM-Ca2+. The main conductance level varied from patch to patch and the presence (or absence) of additional conductance levels also varied. 2. The channels showed large open channel noise and experiments with 300 mM-NaCl in the patch pipette substantially increased the open channel noise. The appearance of detectable step-like transitions within this noise strongly suggested the existence of closely spaced discrete levels. 3. Removal of divalent cations from the external solution increased the unitary channel conductance. Altering the main permeant ion in divalent-free solutions gave the following conductance sequence: K+ (93 pS) greater than Cs+ (61 pS) greater than Na+ (51 pS) greater than Li+(23 pS). 4. Replacement of Na+ by Cs+ in the external solution considerably reduced the current evoked by ACh in whole-cell recordings and the channel-opening frequency in outside-out patches. 5. The kinetic properties of channels activated by ACh and 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP) were also studied. At low concentrations of ACh and DMPP the gap distributions were complex and best fitted by the sum of four exponential components. Individual activations (bursts) were interrupted by the two shortest closed periods the briefer of which had time constants of 36 microseconds for ACh and 67 microseconds for DMPP. 6. The distribution of burst lengths had two components for each agonist, each component making up about 50% of the total area under the distribution. For ACh, the time constant of the longer component (12.2 ms) was similar to the decay time constant of excitatory postsynaptic potentials (EPSCs) at similar temperature and potential. For DMPP the time constant of the longer component was 17.6 ms. 7. The relative number of brief gaps per long burst was much larger for ACh than for DMPP. Therefore the corrected mean open time for ACh (0.86 ms) was much shorter than that for DMPP (2.3 ms). 8. In terms of receptor mechanism, the values of the channel opening equilibrium constant (beta/alpha) estimated from these numbers (ACh, 23; DMPP, 25) suggest that both agonists are efficaceous. 9. DMPP is a potent blocker of the channel with an equilibrium dissociation constant (KB) of around 50 microM and blockage gaps of around 1 ms duration. ACh also blocks the channel but with a higher KB of around 470 microM.(ABSTRACT TRUNCATED AT 400 WORDS)
Mathie
A
A
Department of Pharmacology, University College London.
Cull-Candy
S G
SG
Colquhoun
D
D
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D004246:Dimethylphenylpiperazinium Iodide / Q000494:pharmacology; D005260:Female; D005728:Ganglia, Sympathetic / Q000502:physiology*; D015640:Ion Channel Gating / Q000187:drug effects; D007473:Ion Channels / Q000502:physiology*; D008564:Membrane Potentials / Q000187:drug effects; D051381:Rats; D011919:Rats, Inbred Strains; D011978:Receptors, Nicotinic / Q000187:drug effects / Q000502:physiology*; D012964:Sodium / Q000494:pharmacology; D013997:Time Factors
D007473:Ion Channels; D011978:Receptors, Nicotinic; D004246:Dimethylphenylpiperazinium Iodide; D012964:Sodium; D000109:Acetylcholine