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PF4, a FMRFamide-related peptide, gates low-conductance Cl(-) channels in Ascaris suum
Jenny Purcell1, Alan P Robertson, David P Thompson
1Department of Preclinical Veterinary Science, University of Edinburgh, Summerhall, UK.
Abstract:
Here we describe the actions of the peptide Lys-Pro-Asn-Phe-Ile-Arg-Phe-NH(2), or PF4, on inside-out membrane patches (n=164), recorded from vesicles derived from Ascaris suum somatic muscle cells. We observed numerous, small-amplitude Cl(-) channels in the membrane patches. The conductance of the Cl(-) channels ranged from 1.09 to 7.07 pS, the open probability (P(open)) ranged from 0.047+/-0.015 (mean+/-S.E.M.) at 0 microM PF4 to 0.156+/-0.026 at 0.1 microM PF4. The channel mean open time was more variable and prolonged at negative potentials than when the membrane patch was clamped at positive potentials: at 0.03 microM PF4, the mean open time (+/-S.E.M) at -80 mV was 522+/-333 ms; at+80 mV, it was 25+/-7 ms. When patches were isolated from the parent vesicle, there were no changes in channel characteristics, suggesting that the channels function without the involvement of cytoplasmic components. Similarly, the channel characteristics were not affected by the G-protein inhibitor, guanosine-5'-O-(2-thiodiphosphate), indicating that the ion channels do not require a G-protein to function. These data indicate that the PF4-activated Cl(-) channels function independently of intracellular signal transducers and are, therefore, directly gated by PF4.
Insights
The peptide PF4 directly activates chloride channels in Ascaris suum muscle cells. These channels function independently of cytoplasmic factors and G-proteins, indicating a direct gating mechanism by PF4.
Area of Science:
- Biophysics
- Molecular Biology
- Cell Physiology
Background:
- Chloride channels play crucial roles in cellular functions.
- The peptide PF4's role in ion channel regulation is not fully understood.
Purpose of the Study:
- To investigate the effect of the peptide PF4 on chloride channels in Ascaris suum somatic muscle cells.
- To determine the gating mechanism of PF4-activated chloride channels.
Main Methods:
- Utilized inside-out membrane patch-clamp recordings from Ascaris suum somatic muscle cell-derived vesicles.
- Characterized chloride channel properties including conductance, open probability, and mean open time under varying PF4 concentrations and membrane potentials.
- Tested the involvement of cytoplasmic components and G-proteins using isolated patches and a G-protein inhibitor.
Main Results:
- Observed numerous small-amplitude chloride channels with conductances ranging from 1.09 to 7.07 pS.
- PF4 increased channel open probability from 0.047 to 0.156 with increasing concentration.
- Channel open time was dependent on membrane potential and PF4 concentration, with longer durations at negative potentials.
- Channel characteristics remained unchanged in isolated patches and in the presence of a G-protein inhibitor.
Conclusions:
- PF4 directly gates chloride channels in Ascaris suum somatic muscle cells.
- These chloride channels function independently of cytoplasmic factors and G-protein signaling pathways.
- The findings suggest a direct interaction between PF4 and the chloride channel protein.