Abstract
Iptakalim, a novel cardiovascular ATP-sensitive K+ (KATP) channel opener, exerts neuroprotective effects on dopaminergic (DA) neurons against metabolic stress-induced neurotoxicity, but the mechanisms are largely unknown. Here, we examined the effects of iptakalim on functional KATP channels in the plasma membrane (pm) and mitochondrial membrane using patch-clamp and fluorescence-imaging techniques. In identified DA neurons acutely dissociated from rat substantia nigra pars compacta (SNc), both the mitochondrial metabolic inhibitor rotenone and the sulfonylurea receptor subtype (SUR) 1-selective KATP channel opener (KCO) diazoxide induced neuronal hyperpolarization and abolished action potential firing, but the SUR2B-selective KCO cromakalim exerted little effect, suggesting that functional KATP channels in rat SNc DA neurons are mainly composed of SUR1. Immunocytochemical staining showed a SUR1-rather than a SUR2B-positive reaction in most dissociated DA neurons. At concentrations between 3 and 300 μM, iptakalim failed to hyperpolarize DA neurons; however, 300 μM iptakalim increased neuronal firing. In addition, iptakalim restored DA neuronal firing during rotenone-induced hyperpolarization and suppressed rotenone-induced outward current, suggesting that high concentrations of iptakalim close neuronal KATP channels. Furthermore, in human embryonic kidney 293 cells, iptakalim (300-500 μM) closed diazoxide-induced Kir6.2/SUR1 KATP channels, which were heterologously expressed. In rhodamine-123-preloaded DA neurons, iptakalim neither depolarized mitochondrial membrane nor prevented rotenone-induced mitochondrial depolarization. These data indicate that iptakalim is not a KATP channel opener in rat SNc DA neurons; instead, iptakalim is a pm-KATP channel closer at high concentrations. These effects of iptakalim stimulate further pharmacological investigation and the development of possible therapeutic applications.
Footnotes
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This work was supported by the Women's Board Foundation of the Barrow Neurological Institute.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.106.106286.
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ABBREVIATIONS: PD, Parkinson's disease; pm, plasma membrane; mito, mitochondrial; DA, dopaminergic; SNc, substantia nigra pars compacta; SUR, sulfonylurea; HEK, human embryonic kidney; TH, tyrosine hydroxylase; PBS, phosphate-buffered saline; Rh-123, rhodamine-123; P1075, N-cyano-N″-(1,1,-dimethylpropyl)-N″-3-pyridylguanidine; DZX, diazoxide; ACSF, artificial cerebrospinal fluid.
- Received April 14, 2006.
- Accepted July 10, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
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