Ca Entry Research Articles

MILD HYPOXIA/ACIDOSIS INCREASE THE FREQUENCY OF CALCIUM OSCILLATIONS TO AUGMENT SECRETORY ACTIVITY IN RAT CAROTID BODY CHEMORECEPTORS

In carotid body (CB) chemoreceptors (Type 1/gloms cells), severe levels of hypoxia (<1%O2) and acidosis (pH 5.0–6.5) are well known to produce strong depolarization and sustained elevation of intracellular [Ca], which enhances transmitter secretion. We studied the effects of mild and moderate levels of hypoxia (2–5%O2) and acidosis (pH7.0–7.3) on cell membrane potential (Em), Ca signaling and catecholamine secretion in Type 1 cells isolated from 3–4 week-old rats. Spontaneous Ca oscillations of varying frequency and amplitude were observed under normoxic conditions in many individual Type 1 cells in cluster. The frequency and amplitude of Ca oscillations were perfusion flow- and temperature-dependent. Verapamil (an inhibitor of both T- and L- type Ca channels), 2-APB (inhibitor of IP3 receptor and store-operated Ca entry) and removal of external Ca abolished Ca oscillations. Mild/moderate hypoxia (2–5%O2) produced low level depolarization (<12 mV) and increased the frequency and amplitude of Ca oscillations. Mild/moderate acidosis (pH 7.0–7.2) also produced small depolarization (<15 mV) and increased the frequency and amplitude of Ca oscillations. These effects of hypoxia and acidosis were reduced to varying degrees by Ca channel inhibitors, but were not inhibited by amiloride. In CB slices, both hypoxia and acidosis caused small increases in catecholamine secretion as assessed by amperometry. These findings show that mild hypoxia and acidosis increase the frequency and amplitude of Ca oscillations, thereby augmenting the secretory activity of the CB. Support or Funding Information This work was supported by funding from NIH Ca oscillations in gloms cells: effect of 2-APB This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Pharmacological and Ischemic Preconditioning Up-Regulate the Expression of Socs in Adult Cardiac Myocytes: Its Physiological Significance

Voltage- and SOCS channels are the major routes of Ca(2+) entry in mammalian cells and a negative reciprocal interaction between these channels has been described (Doltmetsch et al., 2010). We have previously reported (Gonzalez et al., 2010) that preconditioning in heart leads to down-regulation of Cav1.2 channels. In the present experiments we examined the changes that take place by preconditioning in the expression and function of STIM1 and Orai1, major components of SOCS in heart. We measured membrane currents with the whole-cell patch-clamp technique. To assess the effect of preconditioning on SOCS, protein expression levels of STIM1 and Orai1 were measured by Western blotting and confocal microscopy, and intracellular Ca(2+) measurements was quantitated by Fura-2. Diazoxide (100 uM), was used to induce pharmacological preconditioning. Ischemic preconditioning was produced by brief periods of ischemia. Depletion of internal stores was achieved with thapsigargin. Both forms of preconditioning greatly reduced infaction areas relative to controls. Store depletion generated Ca(2+) selective currents with strong inward rectification. Preconditioning almost completely suppressed these currents, an effect that was almost blocked by 5HD a selective mitoKATP channel blocker and by intracellular BAPTA. And as assessed by Fura-2 this influx was also blocked by preconditioning. In contrast, the expression at the protein level of both STIM1 and Orai1 was strongly up-regulated by preconditioning and confocal microscopy revealed a higher density of Orai1 channels at the surface membrane. Our results indicate that while the expression of protein components of SOCS in heart cells (STIM1 and Orai1) are up-regulated by preconditioning, influx of Ca(2+) through SOCS is severely impaired by preconditioning, probably by a slow Ca(2+)-dependent inactivation process. Supported by CONACYT, grant 284053 to JS.

Activation of store – operated Ca(2+) entry in cisplatin resistant leukemic cells after treatment with photoexcited fullerene C(60) and cisplatin

Activation of store – operated Ca(2+) entry in cisplatin resistant leukemic cells after treatment with photoexcited fullerene C(60) and cisplatin

Ellagic Acid exerts anti‐inflammatory actions via disruption of store‐operated Ca entry (SOCE) pathway activators and coupling mediators[Office1

Pharmacological agents that can modulate Ca2+ homeostasis are being extensively explored as potential anti‐inflammatory agents. Ellagic acid (EA), a polyphenolic compound found in many fruits and plant extracts, has been known to possess anti‐inflammatory properties. However, the mechanism of action behind its anti‐inflammatory action is unclear. We studied the effects of EA on Ca2+ handling, in particular, store‐operated Ca2+ entry (SOCE), a process critical to proper T cell function. EA is thought to modulate Ca2+ homeostasis by enhancing sarcoplasmic endoplasmic reticulum ATPase pump (SERCA) activity, however effects of EA on Ca2+ signaling is not clearly understood. We demonstrate that acute addition of EA induced Ca2+ release (EC50= 63 μM). A 12 h incubation of Jurkat T cells with EA decreased the ATP‐induced Ca2+ release and diminished the SOCE‐mediated Ca2+ influx in a dose dependent manner (IC50= 23 μM). The inhibition of SOCE was also confirmed by reduced Mn2+ entry rates in the EA‐treated cells. The two important components of SOCE are the endoplasmic reticulum (ER)‐Ca2+ sensor Stromal Interaction Molecule (STIM), and highly‐Ca2+ selective plasma membrane (PM) Orai channels. EA treatment attenuated the ATP‐induced Ca2+ influx in EA‐treated HEK293 cells transiently transfected with SOCE channel Orai1‐myc and ER‐sensor stromal interaction molecule (STIM1) (HEKSTIM/Orai). Furthermore, EA treatment interfered with the Orai1 and STIM1 coupling by disrupting STIM1 puncta formation in the HEKSTIM/Orai cells. Since expression of certain cytokines is directly dependent on Ca2+ influx, we measured the levels of IL‐2 and INFγ in EA‐treated Jurkat cells. Indeed, EA‐treated resulted in a significant decrease in IL‐2 and INFγ expression. The activity of nuclear factor of activated T‐cell (NFAT), a transcriptional factor has been shown to be dependent on SOCE mediated Ca2+ influx. Prolong exposure to EA decreased the translocation of nuclear factor of activated T‐cell in stimulated T cells. Hence, by inhibiting SOCE mediated Ca2+ influx, EA decreased downstream activation of pro‐inflammatory mediators. These observations suggest a novel target to EA‐mediated effects and provide insight into the mechanisms underlying EA mediated anti‐inflammatory effects.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Transient Receptor Potential Canonical Channel Blockers Improve Ventricular Contractile Functions After Ischemia/Reperfusion in a Langendorff-perfused Mouse Heart Model.

Reperfusion of ischemic myocardium is accompanied by intracellular Ca overload, leading to cardiac dysfunction. However, the mechanisms underlying intracellular Ca overload have yet to be fully elucidated. The mechanism may involve the activation of store-operated Ca entry, which is primarily mediated through the transient receptor potential canonical (TRPC) channels. This study was undertaken to examine the possible involvement of TRPC channels in the development of contractile dysfunction associated with reperfusion of ischemic myocardium using a mouse heart model. The functional expression of TRPC channels was confirmed in mouse ventricular myocytes using immunocytochemistry, Western blotting, and patch-clamp experiments. The left ventricular functions were assessed by measuring left ventricular end-diastolic pressure, left ventricular developed pressure, and its first derivatives in a Langendorff-perfused mouse heart subjected to 30 minutes of normothermic (37°C) global ischemia followed by 60 minutes of reperfusion. Under control conditions, left ventricular functions were deteriorated during reperfusion, which was significantly ameliorated by administration of the TRPC channel blockers 2-aminoethoxydiphenyl borate and La during initial 5 minutes of reperfusion. Our findings suggest that TRPC channels are involved in mediating contractile dysfunction during reperfusion of ischemic myocardium and detect TRPC channels as a potential therapeutic target for preventing myocardial ischemia/reperfusion injury.

Extracellular pH Regulates the Entry of Calcium into Jurkat T-cells

The entry of calcium into blood cells is a key event in the launching of multiple transcription programs and in the development of immune response, proliferation, growth and cell differentiation. The TRPchannel superfamily serves as the main conductor of calcium ions in blood cells, most of the regulatory elements of which remain unknown. In the current work carried out on Jurkat cells using the patch-clamp method, we showed that the activity of TRPV5 and TRPV6 calcium channels (transient receptor potential vanilloid) of the TRP superfamily depends on the pH of the environment. Results of measurements of the intracellular Ca2+ level using a fluorescent Ca2+-specific Fluo-4 probe demonstrated that the entry of Ca2+ into Jurkat cells is also regulated by extracellular pH. The obtained data indicate that changes in the pH of the environment affecting the activity of the TRPV5 and TRPV6 calcium channels can regulate the entry and level of intracellular Ca2+ in Jurkat cells. It is suggested that such a mechanism for responding to changes in the acid–base balance is necessary for the cells of the immune system to trigger protective reactions, including the areas of development of various inflammations.

Channeling Vision: CaV1.4—A Critical Link in Retinal Signal Transmission

Voltage-gated calcium channels (VGCC) are key to many biological functions. Entry of Ca2+ into cells is essential for initiating or modulating important processes such as secretion, cell motility, and gene transcription. In the retina and other neural tissues, one of the major roles of Ca2+-entry is to stimulate or regulate exocytosis of synaptic vesicles, without which synaptic transmission is impaired. This review will address the special properties of one L-type VGCC, CaV1.4, with particular emphasis on its role in transmission of visual signals from rod and cone photoreceptors (hereafter called “photoreceptors,” to the exclusion of intrinsically photoreceptive retinal ganglion cells) to the second-order retinal neurons, and the pathological effects of mutations in the CACNA1F gene which codes for the pore-forming α1F subunit of CaV1.4.

Ups and downs of calcium in the heart.

Contraction and relaxation of the heart result from cyclical changes of intracellular Ca2+ concentration ([Ca2+ ]i ). The entry of Ca2+ into the cell via the L-type Ca2+ current leads to the release of more from the sarcoplasmic reticulum (SR). Compared to other regulatory mechanisms such as phosphorylation, Ca2+ signalling is very rapid. However, since Ca2+ cannot be destroyed, Ca2+ signalling can only be controlled by pumping across membranes. In the steady state, on each beat, the amount of Ca2+ released from the SR must equal that taken back and influx and efflux across the sarcolemma must be equal. Any imbalance in these fluxes will result in a change of SR Ca2+ content and this provides a mechanism for regulation of SR Ca2+ content. These flux balance considerations also explain why simply potentiating Ca2+ release from the SR has no maintained effect on the amplitude of the Ca2+ transient. A low diastolic [Ca2+ ]i is essential for cardiac relaxation, but the factors that control diastolic [Ca2+ ]i are poorly understood. Recent work suggests that flux balance is also important here. In particular, decreasing SR function decreases the amplitude of the systolic Ca2+ transient and the resulting decrease of Ca2+ efflux results in an increase of diastolic [Ca2+ ]i to maintain total efflux.

Increased calcium uptake and improved trabecular bone properties in intestinal alkaline phosphatase knockout mice.

Previous studies have demonstrated a negative correlation between intestinal alkaline phosphatase (IAP) activity and calcium (Ca) absorption in the gut, as IAP acts as a protective mechanism inhibiting high Ca entry into enterocytes, preventing Ca overload. Here we evaluated Ca absorption and bone properties in knockout mice (KO) completely devoid of duodenal IAP (Akp3 -/- mice). Female C57BL/6 control mice (WT, n=7) and KO mice (n=10) were used to determine Ca absorption in vivo and by in situ isolated duodenal loops followed by histomorphometric analysis of duodenal villi and crypts. Bone mineral density, morphometry, histomorphometry and trabecular connectivity and biomechanical properties were measured on bones. We observed mild atrophy of the villi with lower absorption surface and a significantly higher Ca uptake in KO mice. While no changes were seen in cortical bone, we found better trabecular connectivity and biomechanical properties in the femurs of KO mice compared to WT mice. Our data indicate that IAP KO mice display higher intestinal Ca uptake, which over time appears to correlate with a positive effect on the biomechanical properties of trabecular bone.

Altered mitochondrial function, capacitative calcium entry and contractions in the aorta of hypertensive rats.

It has been suggested that Ca entry through store-operated Ca channels (SOCs) is regulated by a dynamic interplay between the endoplasmic reticulum Ca stores and the mitochondria. These relationships drive the activation and inactivation of SOCs, yet it remains unclear whether this regulation of SOCs by mitochondria is altered in the aorta of spontaneously hypertensive rats (SHRs). We performed a thorough study of the mitochondrial membrane potential, the ability of mitochondria to deal with cytosolic Ca, capacitative Ca entry (CCE), and stromal interaction molecule 1 (STIM1) and calcium release-activated calcium modulator 1 (orai1) protein expression, as well as the contractile capacity of aortic rings, in normotensive Wistar Kyoto rats (WKYs) and SHRs. Changes were observed in aortic tissue and cultured vascular smooth muscle cells isolated from SHRs relative to WKYs, including more depolarized mitochondria, stronger CCE upon the addition of Ca, larger cytosolic Ca transients (cytosolic Ca concentration) or aortic ring contraction elicited by endoplasmic reticulum depletion and a significant increase in STIM1 protein expression but not of orai1. These results suggest that the impaired Ca buffering capacity of partially depolarized mitochondria dysregulates CCE, leading to overfilling of the endoplasmic reticulum Ca store through enhanced STIM1/orai1 interactions and an increase in aorta contractions in SHRs. Thus, understanding the implications of the alterations to STIM1/orai1, and their relationship to mitochondria, may aid drug development and therapeutic strategies to treat hypertension, as well as its long-term sequelae in poorly controlled patients.

P1092ROS-dependent store-operated Ca entry does not increase diastolic Ca in isolated ventricular myocytes

P1092ROS-dependent store-operated Ca entry does not increase diastolic Ca in isolated ventricular myocytes

Systolic [Ca2+ ]i regulates diastolic levels in rat ventricular myocytes.

Key points For the heart to function as a pump, intracellular calcium concentration ([Ca2+]i) must increase during systole to activate contraction and then fall, during diastole, to allow the myofilaments to relax and the heart to refill with blood.The present study investigates the control of diastolic [Ca2+]i in rat ventricular myocytes.We show that diastolic [Ca2+]i is increased by manoeuvres that decrease sarcoplasmic reticulum function. This is accompanied by a decrease of systolic [Ca2+]i such that the time‐averaged [Ca2+]i remains constant.We report that diastolic [Ca2+]i is controlled by the balance between Ca2+ entry and Ca2+ efflux during systole.The results of the present study identify a novel mechanism by which changes of the amplitude of the systolic Ca transient control diastolic [Ca2+]i. The intracellular Ca concentration ([Ca2+]i) must be sufficently low in diastole so that the ventricle is relaxed and can refill with blood. Interference with this will impair relaxation. The factors responsible for regulation of diastolic [Ca2+]i, in particular the relative roles of the sarcoplasmic reticulum (SR) and surface membrane, are unclear. We investigated the effects on diastolic [Ca2+]i that result from the changes of Ca cycling known to occur in heart failure. Experiments were performed using Fluo‐3 in voltage clamped rat ventricular myocytes. Increasing stimulation frequency increased diastolic [Ca2+]i. This increase of [Ca2+]i was larger when SR function was impaired either by making the ryanodine receptor leaky (with caffeine or ryanodine) or by decreasing sarco/endoplasmic reticulum Ca‐ATPase activity with thapsigargin. The increase of diastolic [Ca2+]i produced by interfering with the SR was accompanied by a decrease of the amplitude of the systolic Ca transient, such that there was no change of time‐averaged [Ca2+]i. Time‐averaged [Ca2+]i was increased by β‐adrenergic stimulation with isoprenaline and increased in a saturating manner with increased stimulation frequency; average [Ca2+]i was a linear function of Ca entry per unit time. Diastolic and time‐averaged [Ca2+]i were decreased by decreasing the L‐type Ca current (with 50 μm cadmium chloride). We conclude that diastolic [Ca2+]i is controlled by the balance between Ca entry and efflux during systole. Furthermore, manoeuvres that decrease the amplitude of the Ca transient (without decreasing Ca influx) will therefore increase diastolic [Ca2+]i. This identifies a novel mechanism by which changes of the amplitude of the systolic Ca transient control diastolic [Ca2+]i.

Calcium and Excitation-Contraction Coupling in the Heart.

Cardiac contractility is regulated by changes in intracellular Ca concentration ([Ca2+]i). Normal function requires that [Ca2+]i be sufficiently high in systole and low in diastole. Much of the Ca needed for contraction comes from the sarcoplasmic reticulum and is released by the process of calcium-induced calcium release. The factors that regulate and fine-tune the initiation and termination of release are reviewed. The precise control of intracellular Ca cycling depends on the relationships between the various channels and pumps that are involved. We consider 2 aspects: (1) structural coupling: the transporters are organized within the dyad, linking the transverse tubule and sarcoplasmic reticulum and ensuring close proximity of Ca entry to sites of release. (2) Functional coupling: where the fluxes across all membranes must be balanced such that, in the steady state, Ca influx equals Ca efflux on every beat. The remainder of the review considers specific aspects of Ca signaling, including the role of Ca buffers, mitochondria, Ca leak, and regulation of diastolic [Ca2+]i.

Inhibition of glutamate release by cilnidipine in rat cerebrocortical nerve terminals (synaptosomes).

Cilnidipine is an antihypertensive drug that was reported to have a neuroprotective profile. The present study aimed to investigate the effect of cilnidipine on the 4-aminopyridine (4-AP)-induced glutamate release in the rat cerebral cortex using isolated nerve terminals (synaptosomes). Cilnidipine reduced the release of glutamate release induced by 4-AP in a concentration-dependent manner. This inhibitory effect was associated with a reduction in the 4-AP-induced intrasynaptosomal Ca concentration elevation and was not because of an alteration of the synaptosomal membrane potential. The inhibition of glutamate release by cilnidipine was markedly reduced or eliminated in the presence of the Cav2.2 (N-type) and Cav2.1 (P/Q-type) channel blocker ω-conotoxin MVIIC and the protein kinase A inhibitor H89. The intracellular Ca-release inhibitors dantrolene and CGP37157, the mitogen-activated protein kinase inhibitor PD98059 or the protein kinase C inhibitor GF109203X failed to affect the action of cilnidipine. These results suggest that cilnidipine inhibits glutamate release from rat cortical synaptosomes through the suppression of presynaptic voltage-dependent Ca entry and protein kinase A activity.

Caffeine Inhibits Fluid Secretion by Interlobular Ducts From Guinea Pig Pancreas.

Caffeine is contained in coffee, tea, and numerous beverages and foods. We examined the direct effects of caffeine on the physiological function of pancreatic duct cells by using interlobular duct segments isolated from guinea pig pancreas. The rate of fluid secretion was continuously measured by monitoring the luminal volume of isolated duct segments. Changes in intracellular Ca concentration ([Ca]i) were estimated by microfluorometry in ducts loaded with Fura-2. Both secretin-stimulated and acetylcholine (ACh)-stimulated fluid secretions were substantially and reversibly inhibited by relatively low concentrations of caffeine as low as 0.03 mM relevant to blood levels after ingestion of caffeine-containing beverages. Caffeine inhibited ACh-induced elevation of [Ca]i and secretin-induced fluctuation of [Ca]i. Caffeine abolished thapsigargin-induced intracellular Ca release but did not affect the entry of extracellular Ca. Caffeine (0.05 mM) abolished ethanol (1 mM)-induced fluid hypersecretion in secretin-stimulated pancreatic duct. Low concentrations of caffeine directly inhibit pancreatic ductal fluid secretion stimulated by secretin or ACh and also ethanol-induced fluid hypersecretion. The inhibition by caffeine seems to be mediated by the blockade of intracellular Ca mobilization. Daily intake of caffeine may reduce the volume of pancreatic juice secretion.

Mechanisms of electromechanical and electrochemical coupling in olfactory cilia of the frog (Rana temporaria)

The mechanisms of electromechanical and electrochemical coupling in olfactory cilia of the frog (Rana temporaria) have been investigated. High-resolution optical television microscopy of live tissue and pharmacological analysis have been used to reveal the regulation of the motility of olfactory cilia in the absence of odorants; the entry of Ca2+ ions mediated by three types of ion channels (mechanosensitive, cyclic nucleotide gated, and voltage-gated) was shown to determine the motility of cilia. Stimulation of the olfactory adenylate cyclase by movements of the cilia in the absence of odors has been demonstrated and the regulation of cilia motility by membrane potential has been revealed. Membrane potential can affect olfactory acuity and the ability to perceive weak olfactory stimuli in the absence of adequate stimulation.

Effect of piracetam on Ca2+-induced K+ efflux from sickle cells

We have therefore studied the in vitro effect of piracetam on Ca 2+ -activated K + ( 86 Rb) efflux from sickle cells. We have aslo tested the protective effect of piracetam on the deformability of sickle cells when entry of Ca 2+ was induced by repetitive cycles of oxygenation-deoxygenation for 15 hours

Novel function of α1D L-type calcium channel in the atria

Ca entry through atrial L-type Calcium channels (α1C and α1D) play an important role in muscular contraction, regulation of gene expression, and release of hormones including atrial natriuretic peptide (ANP), and brain natriuretic peptide (BNP). α1D Ca channel is exclusively expressed in atria, and has been shown to play a key role in the pathogenesis of atrial fibrillation. Recent data have shown that the small conductance calcium-activated potassium channel, SK4 is also atrial specific and also contributes prominently to the secretion of ANP and BNP. However, its functional role in the heart is still poorly understood. Here we used α1D gene heterozygous (α1D+/-) mice and HL-1 cells to determine the functional contribution of SK4 channels to α1D–dependent regulation of ANP and BNP secretion in response to endothelin (ET), and/or mechanical stretch. Immunoprecipitation with α1D specific antibody and western blotting with SK4 specific antibody on the immuno-precipitated protein complex showed a band at 50 KDa confirming the presence of SK4 in the complex and provided evidence of interaction between SK4 and α1D channels. Using RT-PCR, we observed a 2.9 fold decrease in expression of Cacna1d (gene encoding α1D) mRNA in atria from α1D+/−mice. The decrease in α1D mRNA corresponded with a 4.2 fold decrease in Kcnn4 (gene encoding SK4) mRNA from α1D+/− mice. These changes were paralleled with a 77% decrease in BNP serum levels from α1D+/− mice. When α1D was knocked down in HL-1cardiomyocytes using CRISPR/Cas9 technology, a 97% decrease in secreted BNP was observed even in cells subjected to stretch and endothelin. In conclusion, our data are first to show that α1D Ca and SK4 channels are coupled in the atria, and that deletion of α1D leads to decreased SK4 mRNA and BNP secretion providing evidence for a novel role of α1D in atrial endocrine function. Elucidating the regulatory factors that underlie the secretory function of atria will identify novel therapeutic targets for treatment and prevention of cardiac arrhythmias such as atrial fibrillation.

The Inositol Trisphosphate/Calcium Signaling Pathway in Health and Disease.

Many cellular functions are regulated by calcium (Ca(2+)) signals that are generated by different signaling pathways. One of these is the inositol 1,4,5-trisphosphate/calcium (InsP3/Ca(2+)) signaling pathway that operates through either primary or modulatory mechanisms. In its primary role, it generates the Ca(2+) that acts directly to control processes such as metabolism, secretion, fertilization, proliferation, and smooth muscle contraction. Its modulatory role occurs in excitable cells where it modulates the primary Ca(2+) signal generated by the entry of Ca(2+) through voltage-operated channels that releases Ca(2+) from ryanodine receptors (RYRs) on the internal stores. In carrying out this modulatory role, the InsP3/Ca(2+) signaling pathway induces subtle changes in the generation and function of the voltage-dependent primary Ca(2+) signal. Changes in the nature of both the primary and modulatory roles of InsP3/Ca(2+) signaling are a contributory factor responsible for the onset of a large number human diseases.

Relaxant Effects of the Selective Estrogen Receptor Modulator, Bazedoxifene, and Estrogen Receptor Agonists in Isolated Rabbit Basilar Artery.

We have previously shown that the selective estrogen receptor modulator, bazedoxifene, improves the consequences of ischemic stroke. Now we aimed to characterize the effects and mechanisms of action of bazedoxifene in cerebral arteries. Male rabbit isolated basilar arteries were used for isometric tension recording and quantitative polymerase chain reaction. Bazedoxifene relaxed cerebral arteries, as 17-β-estradiol, 4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol [estrogen receptor (ER) α agonist], and G1 [G protein-coupled ER (GPER) agonist] did it (4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol > bazedoxifene = G1 > 17-β-estradiol). 2,3-Bis(4-hydroxyphenyl)-propionitrile (ERβ agonist) had no effect. Expression profile of genes encoding for ERα (ESR1), ERβ (ESR2), and GPER was GPER > ESR1 > ESR2. As to the endothelial mechanisms, endothelium removal, N-nitro-L-arginine methyl ester, and indomethacin, did not modify the relaxant responses to bazedoxifene. As to the K channels, both a high-K medium and the Kv blocker, 4-aminopyridine, inhibited the bazedoxifene-induced relaxations, whereas tetraethylammonium (nonselective K channel blocker), glibenclamide (selective KATP blocker) or iberiotoxin (selective KCa blocker) were without effect. Bazedoxifene also inhibited both Ca- and Bay K8644-elicited contractions. Therefore, bazedoxifene induces endothelium-independent relaxations of cerebral arteries through (1) activation of GPER and ERα receptors; (2) increase of K conductance through Kv channels; and (3) inhibition of Ca entry through L-type Ca channels. Such a profile is compatible with the beneficial effects of estrogenic compounds (eg, SERMs) on vascular function and, specifically, that concerning the brain. Therefore, bazedoxifene could be useful in the treatment of cerebral disorders in which the cerebrovascular function is compromised (eg, stroke).