Secretagogue-induced Exocytosis Recruits G Protein-gated K+ Channels to Plasma Membrane in Endocrine Cells

Abstract

Stimulation-regulated fusion of vesicles to the plasma membrane is an essential step for hormone secretion but may also serve for the recruitment of functional proteins to the plasma membrane. While studying the distribution of G protein-gated K ؉ (K G ) channels in the anterior pituitary lobe, we found K G channel subunits Kir3.1 and Kir3.4 localized on the membranes of intracellular dense core vesicles that contained thyrotropin. Stimulation of these thyrotroph cells with thyrotropinreleasing hormone provoked fusion of vesicles to the plasma membrane, increased expression of Kir3.1 and Kir3.4 subunits in the plasma membrane, and markedly enhanced K G currents stimulated by dopamine and somatostatin. These data indicate a novel mechanism for the rapid insertion of functional ion channels into the plasma membrane, which could form a new type of negative feedback control loop for hormone secretion in the endocrine system.The G protein-gated K ϩ (K G ) 1 channel, a member of the inwardly rectifying K ϩ (Kir) channel family, is directly activated by pertussis toxin-sensitive G proteins. This system was first discovered in the muscarinic deceleration of the heartbeat (1). More recently, it is considered to play an essential role in the hormone-mediated inhibitory regulation of neural excitability (1, 2). Electrophysiological studies have revealed that a variety of inhibitory receptors, including M 2 -muscarinic, A 1 -purinergic, D 2 -dopamine, ␣ 2 -adrenergic, serotonin, ␥-aminobutyric acid type B, opioid, and somatostatin receptors in the brain, are coupled to K G channels (3). In endocrine organs such as the anterior pituitary lobe and pancreatic islet, it was also reported that some neurotransmitters including dopamine and somatostatin hyperpolarize the membrane by activating K G channels, which results in the inhibition of hormone secretion (4 -7).The main subunit of K G channels has been cloned from the heart and designated GIRK1/Kir3.1 (8, 9). In the brain and heart, Kir3.1 forms functional K G channels by assembling with other Kir3.0 subunits such as GIRK2/Kir3.2, GIRK3/Kir3.3 (10, 11), and GIRK4/CIR/Kir3.4 (12). Because Kir3.1 mRNA was detected in the anterior pituitary lobe (13) and pancreatic islet (14), this subunit may also contribute to the formation of K G channels in endocrine cells. The anterior pituitary lobe contains several kinds of endocrine cells: lactotrophs, somatotrophs, corticotrophs, and thyrotrophs. Although electrophysiological experiments have shown that somatostatin or dopamine activate K G currents in lactotrophs (4) and also in several cell lines derived from the anterior pituitary lobe, such as GH3 (5) and AtT20 (6), the cellular and subcellular localizations of K G channels in in vivo pituitary endocrine cells have not been examined.In this study, using a polyclonal antibody specific to Kir3.1, we found that this subunit was expressed only in thyrotroph cells and, surprisingly, was localized predominantly on intracellular secretory vesicles. Kir3.4 was co-localized on the vesicles w...

Genes and channels: patch/voltage-clamp analysis and single-cell RT-PCR

“…Voltagegated channels that have been investigated by patch/voltage-clamp analysis and SC-RT-PCR include those with calcium-channel (Plant et al 1998), sodium-channel (O'Dowd andSmith 1996;Vega-Saenz De Miera et al 1997), and potassium-channel subunits. The potassiumchannel subunits that have been investigated by SC-RT-PCR include the Kv1-Kv4 gene family corresponding to the Drosophila potassium-channel genes shaker, shab, shaw, and shal, respectively (Gurantz et al 1996;Baro et al 1997;Massengill et al 1997;Martina et al 1998;Song et al 1998;Seifert et al 1999), and the ATP-sensitive potassium-channel subunits Kir3.1, Kir3.4, Kir4.1, Kir6.1, Kir6.2, SUR1, and SUR2 (Ishii et al 1997;Lee et al 1998Lee et al , 1999Morishige et al 1999;Zawar et al 1999). The combination of whole-cell patch-clamp recording and SC-RT-PCR has also been applied to detect the mRNAs of the calcium-binding proteins, calbindin, parvalbumin, and calretinin, and the neuropeptides, neuropeptide Y, vasoactive intestinal polypeptide, somatostatin, and cholecystokinin, in nonpyramidal cells in acute slices of sensory-motor cortex (Cauli et al 1997).…”

Neuronal activity regulates phosphorylation-dependent surface delivery of G protein-activated inwardly rectifying potassium channels

“…Thus, NMDAR-induced GIRK surface expression may also modulate activity-dependent changes in synaptic strength (synaptic plasticity), such as long-term potentiation, a cellular correlate of learning and memory (15). Given the expected difference in traffic patterns of GIRK channels with different subunit composition (11), it is of interest to note that GIRK channels containing GIRK1 and GIRK4 move from thyroid-stimulating hormone-containing dense core vesicles to the plasma membrane upon vesicle fusion, likely providing feedback regulation of hormone secretion in the anterior pituitary lobe (34).…”

The role of members of the pertussis toxin-sensitive family of G proteins in coupling receptors to the activation of the G protein-gated inwardly rectifying potassium channel

“…Indeed the studies on this channel have become a paradigm of how G ␤␥ can be important in signaling to downstream effectors. Current studies have focused on domains on the channel important for binding G ␤␥ (15)(16)(17)(18)(19)(20), trafficking of the channel complex (21)(22)(23)(24), and the role of anionic phospholipids in regulating channel activity (25)(26)(27)(28)(29).…”