Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Front Pharmacol
2017 May 26;8:302. doi: 10.3389/fphar.2017.00302.
Show Gene links
Show Anatomy links
Sigma-1 Receptor Plays a Negative Modulation on N-type Calcium Channel.
Zhang K
,
Zhao Z
,
Lan L
,
Wei X
,
Wang L
,
Liu X
,
Yan H
,
Zheng J
.
???displayArticle.abstract???
The sigma-1 receptor is a 223 amino acids molecular chaperone with a single transmembrane domain. It is resident to eukaryotic mitochondrial-associated endoplasmic reticulum and plasma membranes. By chaperone-mediated interactions with ion channels, G-protein coupled receptors and cell-signaling molecules, the sigma-1 receptor performs broad physiological and pharmacological functions. Despite sigma-1 receptors have been confirmed to regulate various types of ion channels, the relationship between the sigma-1 receptor and N-type Ca2+ channel is still unclear. Considering both sigma-1 receptors and N-type Ca2+ channels are involved in intracellular calcium homeostasis and neurotransmission, we undertake studies to explore the possible interaction between these two proteins. In the experiment, we confirmed the expression of the sigma-1 receptors and the N-type calcium channels in the cholinergic interneurons (ChIs) in rat striatum by using single-cell reverse transcription-polymerase chain reaction (scRT-PCR) and immunofluorescence staining. N-type Ca2+ currents recorded from ChIs in the brain slice of rat striatum was depressed when sigma-1 receptor agonists (SKF-10047 and Pre-084) were administrated. The inhibition was completely abolished by sigma-1 receptor antagonist (BD-1063). Co-expression of the sigma-1 receptors and the N-type calcium channels in Xenopus oocytes presented a decrease of N-type Ca2+ current amplitude with an increase of sigma-1 receptor expression. SKF-10047 could further depress N-type Ca2+ currents recorded from oocytes. The fluorescence resonance energy transfer (FRET) assays and co-immunoprecipitation (Co-IP) demonstrated that sigma-1 receptors and N-type Ca2+ channels formed a protein complex when they were co-expressed in HEK-293T (Human Embryonic Kidney -293T) cells. Our results revealed that the sigma-1 receptors played a negative modulation on N-type Ca2+ channels. The mechanism for the inhibition of sigma-1 receptors on N-type Ca2+ channels probably involved a chaperone-mediated direct interaction and agonist-induced conformational changes in the receptor-channel complexes on the cell surface.
FIGURE 1. Identification of the expression of N-type Ca2+ channel and sigma-1 receptor in ChIs. (A) An IR-DIC image of a striatal slice illustrating the characteristic appearance of giant cholinergic interneurons (ChIs). (B) Representative photograph of single-cell reverse transcription polymerase chain reaction (scRT-PCR). Experiment products were stained in a 2% agarose gel. Molecular mass markers were shown in the first lane. β-actin and GAPDH were introduced as positive control. The presence of ChAT indicated the aspirated cell was a ChI. (C) Bar plot indicated the co-expression of ChAT, sigma-1 receptor and α1A-C in ChIs by scRT-PCR. ChAT and sigma-1 receptor were in every detected neuron. The α1B subunit of N-type Ca2+ channel was found in 92.1% detected cells. (D) Immunofluorescence of successive paraffin-embedded rat striatum slice observed under 40à objective lens. Left: using the anti-sigma-1 receptor antibody at a dilution of 1:20 and FITC-conjugated affinipure goat anti-rabbit IgG. Middle: Using the anti-ChAT antibody at a dilution of 1:200 and TRITC-conjugated affinipure rabbit anti-goat IgG. Right: the merged picture of the sigma-1 receptor and ChAT.
FIGURE 2. Sigma-1 receptor agonist reduced currents of Ca2+ channel in ChIs. (A) Representative trace of Ca2+ channel current obtained from the rat striatum ChIs in the brain slice. (B) IâV plot of the recorded Ca2+ channel current trace in (A). (CâE) Representative traces of current evoked before and after the bath application of 0.5 μM CTX, 50 μM SKF-10047, and 50 μM Pre-084, respectively. (F) Summarized bar graph to present the effect of CTX (N-type Ca2+ channel blocker), different concentrations of SKF-10047 and Pre-084 (sigma-1 receptor agonists) on Ca2+ currents. (G) Summarized bar graph to present the effect of BD-1063 (sigma-1 receptor antagonist) on the Ca2+ current inhibition induced by SKF-10047. âp < 0.05.
FIGURE 3. Expression of N-type Ca2+ channel and sigma-1 receptor in xenopus oocytes. (A) Upper panel: the current of N-type Ca2+ channel recorded in oocytes after microinjection with N-type Ca2+ channel cRNA. Lower panel: the current was blocked by N-type Ca2+ channel blocker CTX (0.2 μM). (B) I–V plot of the recorded N-type Ca2+ channel current in (A). (C) Schematic Western blot strap of oocytes injected with sigma-1 receptor cRNA (the first lane) and control group (the middle three lanes), which indicated the sigma-1 receptor expressed successfully. The last lane showed the Marker.
FIGURE 4. Currents recorded at different cRNA ratio of sigma-1 receptor to N-type Ca2+ channel. (A) Currents recorded from the oocytes injected with different cRNA ratio. The cRNA concentration ratio of Sigma-1 receptor: N-type Ca2+ channel = 1:1, 0.5:1, 0.25:1, 0.125:1, 0.0625:1, and 0.03125:1, respectively. (B) Summarized bar graph to present the N-type Ca2+ currents decreased with the increase of sigma-1 receptor cRNA injected in oocytes. The current amplitude recorded from oocytes with injection of N-type Ca2+ channel cRNA only was normalized as 100%. Values are Mean ± SEM (n = 18 for 1:1, n = 18 for 0.5:1, n = 17 for 0.25:1, n = 26 for 0.125:1, n = 17 for 0.0625:1, and n = 10 for 0.03125:1 group, respectively. âââp < 0.001, âp < 0.05). (C) The expression levels of the two proteins from oocytes injected with the cRNA ratio of sigma-1 receptor: N-type Ca2+ channel at 0.03125:1, 0.0625:1, 0.125:1, 0.25:1, 0.5:1, and 1:1 from left to right. The blots in the right two lanes showed the protein expression in oocytes of control groups, which were injected with the N-type Ca2+ channel cRNA or the sigma-1 receptor cRNA alone.
FIGURE 5. Effect of sigma-1 receptor agonist and antagonist on N-type Ca2+ channel. (A) Example trace of the inhibition on N-type Ca2+ current induced by 50 μM SKF-10047. The oocyte was injected with a cRNA ratio of sigma 1R: N-type Ca2+ channel = 0.25:1. (B) IâV plot of the recorded N-type Ca2+ current from the oocyte in (A). (C) Summarized bar graph to present the effect of SFK-10047 (sigma-1 receptor agonist) and BD-1063 (sigma-1 receptor antagonist) on N-type Ca2+ currents. Two-way ANOVA analysis, Mean ± SEM, âp < 0.05.
FIGURE 6. Exogenous expression of Sigma-1-Dsred receptor and Cav2.2-GFP in HEK-293T cell line. (A) Fluorescence images of HEK-293T cells transfected with 0.5 μg Sigma-1-Dsred receptor cDNA. (B) Fluorescence images of HEK-293T cells transfected with 0.5 μg Cav2.2-GFP (α1b+β1b+α2δ1subunits) cDNA. (C) Confocal microscopy images of HEK-293T cells with both 0.5 μg Cav2.2-GFP and 0.5 μg Sigma-1-Dsred receptor cDNA. Co-localization was shown in yellow.
FIGURE 7. Proteinâprotein interaction between sigma-1 receptors and N-type Ca2+ channels. (A) HEK-293T cells were transfected with both Cav2.2-GFP (α1b+β1b+α2δ1) and sigma-1-Dsred receptor. Pseudo-color images were obtained under the three filter sets: GFP (a), Dsred (b), and FRET (c). After subtraction of background and bleed-through signals, net FRET (d) was acquired. Normalized FRET (e) values were got by using equation described in section âMaterials and Methods.â Color bars represented relative degree of net FRET and normalized FRET within the cells. (B) Representative cell enlarged from (e). The net FRET value of the point with the arrow was 0.81 ± 0.06 (Mean ± SD). (C) Bar graph of the statistical numbers in 11 experimental fields (203) and 5 negative control fields (0). (D) Co-immunoprecipitation of GFP-Cav2.2 channels and sigma-1-Dsred receptors. Total lysates were prepared from HEK-293T cells. Immunoprecipitated samples were run on the gels and the blots probed with either anti-Cav2.2 antibody or anti-sigma-1 receptor antibody.
Altier,
ORL1 receptor-mediated internalization of N-type calcium channels.
2006, Pubmed
Altier,
ORL1 receptor-mediated internalization of N-type calcium channels.
2006,
Pubmed
Antonini,
Anti-amnesic and neuroprotective actions of the sigma-1 receptor agonist (-)-MR22 in rats with selective cholinergic lesion and amyloid infusion.
2011,
Pubmed
Aydar,
The sigma receptor as a ligand-regulated auxiliary potassium channel subunit.
2002,
Pubmed
,
Xenbase
Carnally,
Demonstration of a direct interaction between sigma-1 receptors and acid-sensing ion channels.
2010,
Pubmed
Cassidy,
Functional exofacially tagged N-type calcium channels elucidate the interaction with auxiliary α2δ-1 subunits.
2014,
Pubmed
Catterall,
Structure and regulation of voltage-gated Ca2+ channels.
2000,
Pubmed
Chu,
Biochemical Pharmacology of the Sigma-1 Receptor.
2016,
Pubmed
Cobos,
Pharmacology and therapeutic potential of sigma(1) receptor ligands.
2008,
Pubmed
Crottès,
Sig1R protein regulates hERG channel expression through a post-translational mechanism in leukemic cells.
2011,
Pubmed
,
Xenbase
Crottès,
The sigma-1 receptor: a regulator of cancer cell electrical plasticity?
2013,
Pubmed
D'Ascenzo,
Electrophysiological and molecular evidence of L-(Cav1), N- (Cav2.2), and R- (Cav2.3) type Ca2+ channels in rat cortical astrocytes.
2004,
Pubmed
Day,
Stimulation of 5-HT(2) receptors in prefrontal pyramidal neurons inhibits Ca(v)1.2 L type Ca(2+) currents via a PLCbeta/IP3/calcineurin signaling cascade.
2002,
Pubmed
Fontanilla,
The hallucinogen N,N-dimethyltryptamine (DMT) is an endogenous sigma-1 receptor regulator.
2009,
Pubmed
Guitart,
Sigma receptors: biology and therapeutic potential.
2004,
Pubmed
Hawkins,
HCN channels contribute to serotonergic modulation of ventral surface chemosensitive neurons and respiratory activity.
2015,
Pubmed
Hayashi,
Intracellular dynamics of sigma-1 receptors (sigma(1) binding sites) in NG108-15 cells.
2003,
Pubmed
Hayashi,
Regulating ankyrin dynamics: Roles of sigma-1 receptors.
2001,
Pubmed
Hayashi,
Sigma-1 receptor chaperones at the ER-mitochondrion interface regulate Ca(2+) signaling and cell survival.
2007,
Pubmed
Johannessen,
Voltage-gated sodium channel modulation by sigma-receptors in cardiac myocytes and heterologous systems.
2009,
Pubmed
Kekuda,
Cloning and functional expression of the human type 1 sigma receptor (hSigmaR1).
1996,
Pubmed
Kinoshita,
Sigma-1 receptor alters the kinetics of Kv1.3 voltage gated potassium channels but not the sensitivity to receptor ligands.
2012,
Pubmed
,
Xenbase
Kisilevsky,
D1 receptors physically interact with N-type calcium channels to regulate channel distribution and dendritic calcium entry.
2008,
Pubmed
Kourrich,
Dynamic interaction between sigma-1 receptor and Kv1.2 shapes neuronal and behavioral responses to cocaine.
2013,
Pubmed
Kourrich,
The sigma-1 receptor: roles in neuronal plasticity and disease.
2012,
Pubmed
Langa,
Generation and phenotypic analysis of sigma receptor type I (sigma 1) knockout mice.
2003,
Pubmed
Lin,
Identification of functionally distinct isoforms of the N-type Ca2+ channel in rat sympathetic ganglia and brain.
1997,
Pubmed
,
Xenbase
Llinás,
Relationship between presynaptic calcium current and postsynaptic potential in squid giant synapse.
1981,
Pubmed
Lu,
σ-1 Receptor agonist SKF10047 inhibits glutamate release in rat cerebral cortex nerve endings.
2012,
Pubmed
Lupardus,
Membrane-delimited coupling between sigma receptors and K+ channels in rat neurohypophysial terminals requires neither G-protein nor ATP.
2000,
Pubmed
,
Xenbase
Luty,
Sigma nonopioid intracellular receptor 1 mutations cause frontotemporal lobar degeneration-motor neuron disease.
2010,
Pubmed
Marriott,
σ-1 receptor at the mitochondrial-associated endoplasmic reticulum membrane is responsible for mitochondrial metabolic regulation.
2012,
Pubmed
Martin,
The effects of morphine- and nalorphine- like drugs in the nondependent and morphine-dependent chronic spinal dog.
1976,
Pubmed
Maurice,
The pharmacology of sigma-1 receptors.
2009,
Pubmed
Maurice,
Sigma(1) (sigma(1)) receptor antagonists represent a new strategy against cocaine addiction and toxicity.
2002,
Pubmed
Maurice,
Neuroprotective and anti-amnesic potentials of sigma (sigma) receptor ligands.
1997,
Pubmed
Mavournin,
The in vivo micronucleus assay in mammalian bone marrow and peripheral blood. A report of the U.S. Environmental Protection Agency Gene-Tox Program.
1990,
Pubmed
Miki,
Activity-dependent neurotrophin signaling underlies developmental switch of Ca2+ channel subtypes mediating neurotransmitter release.
2013,
Pubmed
Mishra,
The sigma-1 receptors are present in monomeric and oligomeric forms in living cells in the presence and absence of ligands.
2015,
Pubmed
Momiyama,
Dopamine D(2)-like receptors selectively block N-type Ca(2+) channels to reduce GABA release onto rat striatal cholinergic interneurones.
2001,
Pubmed
Pal,
Juxtaposition of the steroid binding domain-like I and II regions constitutes a ligand binding site in the sigma-1 receptor.
2008,
Pubmed
Pan,
Sigma-1 receptor antagonism restores injury-induced decrease of voltage-gated Ca2+ current in sensory neurons.
2014,
Pubmed
Quirion,
A proposal for the classification of sigma binding sites.
1992,
Pubmed
Renaudo,
Cancer cell cycle modulated by a functional coupling between sigma-1 receptors and Cl- channels.
2007,
Pubmed
Romieu,
The sigma1 (sigma1) receptor activation is a key step for the reactivation of cocaine conditioned place preference by drug priming.
2004,
Pubmed
Ruscher,
The sigma-1 receptor enhances brain plasticity and functional recovery after experimental stroke.
2011,
Pubmed
Sabeti,
Steroid pregnenolone sulfate enhances NMDA-receptor-independent long-term potentiation at hippocampal CA1 synapses: role for L-type calcium channels and sigma-receptors.
2007,
Pubmed
Schmidt,
Crystal structure of the human σ1 receptor.
2016,
Pubmed
Seth,
Expression pattern of the type 1 sigma receptor in the brain and identity of critical anionic amino acid residues in the ligand-binding domain of the receptor.
2001,
Pubmed
Seth,
Cloning and structural analysis of the cDNA and the gene encoding the murine type 1 sigma receptor.
1997,
Pubmed
Su,
Evidence for sigma opioid receptor: binding of [3H]SKF-10047 to etorphine-inaccessible sites in guinea-pig brain.
1982,
Pubmed
Su,
The Sigma-1 Receptor as a Pluripotent Modulator in Living Systems.
2016,
Pubmed
Su,
The sigma-1 receptor chaperone as an inter-organelle signaling modulator.
2010,
Pubmed
Su,
Understanding the molecular mechanism of sigma-1 receptors: towards a hypothesis that sigma-1 receptors are intracellular amplifiers for signal transduction.
2003,
Pubmed
Tchedre,
Sigma-1 receptor regulation of voltage-gated calcium channels involves a direct interaction.
2008,
Pubmed
Tsai,
Sigma-1 receptor chaperones and diseases.
2009,
Pubmed
Vidal-Torres,
Effects of the selective sigma-1 receptor antagonist S1RA on formalin-induced pain behavior and neurotransmitter release in the spinal cord in rats.
2014,
Pubmed
Xia,
Reliable and global measurement of fluorescence resonance energy transfer using fluorescence microscopes.
2001,
Pubmed
Xu,
Identification of the PGRMC1 protein complex as the putative sigma-2 receptor binding site.
2011,
Pubmed
Yan,
Muscarinic (m2/m4) receptors reduce N- and P-type Ca2+ currents in rat neostriatal cholinergic interneurons through a fast, membrane-delimited, G-protein pathway.
1996,
Pubmed
Yan,
Overexpression of human E46K mutant α-synuclein impairs macroautophagy via inactivation of JNK1-Bcl-2 pathway.
2014,
Pubmed
Zamanillo,
Up-regulation of sigma(1) receptor mRNA in rat brain by a putative atypical antipsychotic and sigma receptor ligand.
2000,
Pubmed
Zhang,
ZC88, a novel N-type calcium channel blocker from 4-amino-piperidine derivatives state-dependent inhibits Cav2.2 calcium channels.
2015,
Pubmed
,
Xenbase
Zhang,
Sigma receptors inhibit high-voltage-activated calcium channels in rat sympathetic and parasympathetic neurons.
2002,
Pubmed
Zhao,
Involvement of HCN Channel in Muscarinic Inhibitory Action on Tonic Firing of Dorsolateral Striatal Cholinergic Interneurons.
2016,
Pubmed
