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Omega-conotoxin blockade distinguishes Ca from Na permeable states in neuronal calcium channels
Abstract:
The blocking properties of the neurotoxic peptide omega-conotoxin GVIA (omega-CgTX), on neuronal Ca channels were investigated. In line with previous reports (Feldman et al. 1987; McCleskey et al. 1987), we found that micromolar concentrations of the toxin block selectively and persistently the high-threshold Ca channels of chick sensory neurons. The block by omega-CgTX could be partially relieved in low [Ca2+]o (less than 1 microM) toxin-free solutions, allowing Na ions to flow through open high-threshold Ca channels. Ca currents through these channels, however, remained permanently blocked on returning to normal Ca2+ toxin-free solutions. Also neurons which were preincubated with omega-CgTX in low Ca2+ (5 mM EGTA) failed to show high-threshold Ca currents during washing with normal Ca2+. Thus, appearance of Na currents through Ca channels in CgTX-pretreated cells was neither a consequence of unbinding of the toxin from its receptor site nor due to an interaction of EGTA with bound omega-CgTX. Na currents in CgTX-pretreated cells could be reversibly suppressed by bath applications of verapamil or by further addition of the toxin. At variance with Ca currents, block of Na currents by omega-CgTX was faster and reversible (KD 0.7 microM). Our data are consistent with the idea that neuronal Ca channels are in different conformational states when permeable to Ca2+ or Na+ ions and that omega-CgTX depresses persistently ion permeation primarily in the Ca-permeable state.
Insights
Omega-conotoxin GVIA (omega-CgTX) persistently blocks high-threshold calcium channels in chick neurons. This neurotoxin affects neuronal Ca channels, influencing ion permeability and conformational states.
Area of Science:
- Neuroscience
- Molecular Biology
- Ion Channel Physiology
Background:
- Neuronal calcium (Ca2+) channels are critical for neurotransmission and neuronal excitability.
- Neurotoxins are valuable tools for probing ion channel function.
- Omega-conotoxin GVIA (omega-CgTX) is a known neurotoxic peptide that interacts with voltage-gated calcium channels.
Purpose of the Study:
- To investigate the blocking properties of omega-conotoxin GVIA (omega-CgTX) on neuronal calcium channels.
- To determine the selectivity and persistence of omega-CgTX block on high-threshold Ca channels.
- To explore the conformational states of neuronal Ca channels in relation to ion permeability.
Main Methods:
- Electrophysiological recordings of ion currents in chick sensory neurons.
- Application of varying concentrations of omega-conotoxin GVIA.
- Manipulation of extracellular calcium ([Ca2+]o) and use of EGTA to assess ion channel block reversibility.
- Pharmacological blockade using verapamil.
Main Results:
- Micromolar concentrations of omega-CgTX selectively and persistently blocked high-threshold Ca channels in chick sensory neurons.
- Partial relief of omega-CgTX block occurred in low [Ca2+]o, allowing Na+ ion flow through Ca channels.
- Ca currents remained permanently blocked upon return to normal Ca2+ conditions, indicating persistent channel modulation.
- Na+ currents in omega-CgTX-pretreated cells were reversibly suppressed by verapamil or additional omega-CgTX, with faster kinetics (KD 0.7 microM).
Conclusions:
- Neuronal Ca channels exist in distinct conformational states when permeable to Ca2+ versus Na+ ions.
- Omega-conotoxin GVIA persistently depresses ion permeation primarily in the Ca2+-permeable state of these channels.
- The findings suggest a model where omega-CgTX stabilizes a non-conducting or altered conducting state of the channel.