Characterization of G-Protein-Gated K⁺Channels Composed of Kir3.2 Subunits in Dopaminergic Neurons of the Substantia Nigra

Abstract

G-protein-gated K+ (KG) channels generate slow inhibitory postsynaptic potentials in the brain. Current opinion suggests that neuronal KG channels are heterotetramers of Kir3.1 and Kir3.2. In substantia nigra (SN), however, mRNA of Kir3.1 does not express, whereas that of Kir3.2 clearly does. Therefore, we have characterized the KG channels containing Kir3.2 subunits in SN using biochemical and immunological techniques. We found that they were composed of only Kir3.2 subunits and did not contain significant amounts of either Kir3.1 or Kir3.3. Furthermore, at least some of the KG channels in SN were assemblies of the splicing variants Kir3. 2a and Kir3.2c. The channels were localized specifically at the postsynaptic membrane on the dendrites of dopaminergic neurons. Kir3. 2c, but not Kir3.2a, could bind a PDZ domain-containing protein, PSD-95. The heterologously expressed KG channels composed of Kir3.2a plus Kir3.2c or Kir3.2a alone were activated by G-protein stimulation, but expression of Kir3.2c alone was not. This study reveals that the Kir3.2 splicing variants play distinct roles in the control of function and localization of some of the KG channels in dopaminergic neurons of SN.

Anchoring proteins confer G protein sensitivity to an inward-rectifier K+ channel through the GK domain

“…It is, however, also possible that unidentified mechanisms might exist that confer functionality to the deaf K G channel. In this study, because Kir3.2c could bind to PSD-95 and SAP97 proteins (Inanobe et al, 1999), we hypothesized that the anchoring proteins might affect the function of Kir3.2c channels, and found that they confer the G protein sensitivity to homomeric Kir3.2c channels. Furthermore, the guanylate kinase (GK) domain of the anchoring proteins was indispensable for the SAP97-induced sensitization of Kir3.2c.…”

“…Thus, they play a key role in generation of slow inhibitory postsynaptic potentials in the brain (Lüscher et al, 1997). We recently found that K G channels in dopaminergic neurons of substantia nigra are localized specifically at the postsynaptic membrane of the dendrites (Inanobe et al, 1999). These K G channels are either heteromultimers of Kir3.2a and Kir3.2c or homomultimers of Kir3.2c alone.…”

“…These K G channels are either heteromultimers of Kir3.2a and Kir3.2c or homomultimers of Kir3.2c alone. However, when the K G channels composed of Kir3.2c alone were expressed in Xenopus oocytes, they did not respond to G protein stimulation at all, although the channel proteins seemed to be generated and transported to the cell membrane as judged from the localization of green fluorescence protein (GFP) tagged to Kir3.2c (Inanobe et al, 1999). Therefore, the homomeric Kir3.2c channel would be produced in vivo but not be functional.…”

Inhibition of G Protein-Activated Inwardly Rectifying K+ Channels by Various Antidepressant Drugs

“…Although GIRK channels in the brain are predominantly GIRK1/2 heteromultimers (Liao et al, 1996), GIRK channels in the substantia nigra and ventral tegmental area are GIRK2 homomultimers (Karschin et al, 1996;Inanobe et al, 1999). To further address the functional relationship between various antidepressants and neuronal GIRK channels, we investigated the effects of the antidepressants on the homomeric channels (Figure 3).…”

“…Four GIRK channel subunits have been identified in mammals (Kubo et al, 1993b;Lesage et al, 1995;Wickman et al, 1997). Neuronal GIRK channels are predominantly heteromultimers composed of GIRK1 and GIRK2 subunits in most brain regions Lesage et al, 1995;Karschin et al, 1996;Liao et al, 1996) or homomultimers composed of GIRK2 subunits in the substantia nigra (Inanobe et al, 1999), whereas atrial GIRK channels are heteromultimers composed of GIRK1 and GIRK4 subunits (Krapivinsky et al, 1995). Various G protein-coupled receptors, such as M 2 muscarinic, a 2 adrenergic, D 2 dopaminergic, 5-HT 1A , opioid, nociceptin/ orphanin FQ and A 1 adenosine receptors, activate GIRK channels (North, 1989;Ikeda et al, 1995Ikeda et al, , 1996Ikeda et al, , 1997 through direct action of G protein bg subunits (Reuveny et al, 1994).…”