Abstract
Recent work has indicated that sigma receptor ligands can modulate potassium channels. However, the only sigma receptor characterized at the molecular level has a novel structure unlike any other receptor known to modulate ion channels. This 26-kDa protein has a hydropathy profile suggestive of a single membrane-spanning domain, with no apparent regions capable of G-protein activation or protein phosphorylation. In the present study patch clamp techniques and photoaffinity labeling were used in DMS-114 cells (a tumor cell line known to express sigma receptors) to investigate the role of the 26-kDa protein in ion channel modulation and probe the mechanism of signal transduction. The sigma receptor ligands N-allylnormetazocine (SKF10047), ditolylguanidine, and (+/-)-2-(N-phenylethyl-N-propyl)-amino-5-hydroxytetralin all inhibited voltage-activated potassium current (IK). Iodoazidococaine (IAC), a high affinity sigma receptor photoprobe, produced a similar inhibition in IK, and when cell homogenates were illuminated in the presence of IAC, a protein with a molecular mass of 26 kDa was covalently labeled. Photolabeling of this protein by IAC was inhibited by SKF10047 with half-maximal effect at 7 microM. SKF10047 also inhibited IK with a similar EC50 (14 microM). Thus, physiological responses to sigma receptor ligands are mediated by a protein with the same molecular weight as the cloned sigma receptor. This indicates that ion channel modulation is indeed mediated by this novel protein. Physiological responses were the same when cells were perfused internally with either guanosine 5'-O-(2-thiodiphosphate) or GTP, indicating that signal transduction is independent of G-proteins. These results demonstrate that ion channels can be modulated by a receptor that does not have seven membrane-spanning domains and does not employ G-proteins. Sigma receptors thus modulate ion channels by a novel transduction mechanism.
Highlights
Sigma receptors are widely distributed in neuronal and nonneuronal tissue and are distinguished by their ability to bind a broad range of chemically unrelated ligands, including (ϩ)
Recent studies in melanotrophs [4] and the neurohypophysis [5] showed that sigma receptor ligands inhibit voltage-activated potassium current (IK), but the signal transduction pathways associated with sigma receptor activation remain unknown
Inhibition of Potassium Current by Sigma Receptor Ligands—DMS-114 cells displayed an outward current in response to depolarizing test pulses under voltage clamp (Fig. 1)
Summary
Sigma receptors are widely distributed in neuronal and nonneuronal tissue and are distinguished by their ability to bind a broad range of chemically unrelated ligands, including (ϩ)-. Inhibition of Potassium Current by Sigma Receptor Ligands—DMS-114 cells displayed an outward current in response to depolarizing test pulses under voltage clamp (Fig. 1). When DMS-114 cells were exposed to the sigma receptor agonist SKF10047, whole-cell IK was inhibited in a concentration-dependent fashion (Fig. 1A).
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