Anchietea pyrifolia A. St.-Hil. as a Cardiovascular-Endowed Species: A Whole-Biological Investigation.

Although leaves of Anchietea salutaris are used in Brazilian traditional medicine, there is no available data in the literature proving its efficacy and safety. Thus, the aim of the study was to perform a meticulous botanical, phytochemical, toxicological, and pharmacological investigation of A. salutaris in Wistar rats. At first, a morphoanatomical characterization of Anchietea pyrifolia leaves and stems was performed. Then, a purified infusion (ethanol-soluble fraction obtained from A. pyrifolia [ESAP]) was obtained followed by its chemical profile elucidation. Furthermore, an acute toxicity test was performed, and the acute and prolonged diuretic and hypotensive effects were also evaluated in Wistar rats. Finally, the vasodilatory responses of ESAP in mesenteric vascular beds were investigated. The main secondary metabolites identified from ESAP were O-glycosylated flavonoids, chlorogenic acids, and phenylpropanoic acid derivatives. ESAP did not promote any toxic effects in female rats nor increased urinary excretion in male rats after a single exposure. However, ESAP significantly reduced renal elimination of sodium, potassium, and chloride after prolonged treatment. An ESAP highest dose promoted significant acute hypotension without affecting blood pressure levels after prolonged use. Furthermore, its cardiovascular effects seem to be related with the calcium-activated potassium channel activation in resistance vessels.

CFTR-mediated anion secretion across intestinal epithelium-like Caco-2 monolayer under PTH stimulation is dependent on intermediate conductance K+ channels.

Parathyroid hormone (PTH), a pleiotropic hormone that maintains mineral homeostasis, is also essential for controlling pH balance and ion transport across renal and intestinal epithelia. Optimization of luminal pH is important for absorption of trace elements, e.g., calcium and phosphorus. We have previously demonstrated that PTH rapidly stimulated electrogenic [Formula: see text] secretion in intestinal epithelial-like Caco-2 monolayers, but the underlying cellular mechanism, contributions of other ions, particularly Cl- and K+, and long-lasting responses are not completely understood. Herein, PTH and forskolin were confirmed to induce anion secretion, which peaked within 1-3 min (early phase), followed by an abrupt decay and plateau that lasted for 60 min (late phase). In both early and late phases, apical membrane capacitance was increased with a decrease in basolateral capacitance after PTH or forskolin exposure. PTH also induced a transient increase in apical conductance with a long-lasting decrease in basolateral conductance. Anion secretion in both phases was reduced under [Formula: see text]-free and/or Cl--free conditions or after exposure to carbonic anhydrase inhibitor (acetazolamide), CFTR inhibitor (CFTRinh-172), Na+/H+ exchanger (NHE)-3 inhibitor (tenapanor), or K+ channel inhibitors (BaCl2, clotrimazole, and TRAM-34; basolateral side), the latter of which suggested that PTH action was dependent on basolateral K+ recycling. Furthermore, early- and late-phase responses to PTH were diminished by inhibitors of PI3K (wortmannin and LY-294002) and PKA (PKI 14-22). In conclusion, PTH requires NHE3 and basolateral K+ channels to induce [Formula: see text] and Cl- secretion, thus explaining how PTH regulated luminal pH balance and pH-dependent absorption of trace minerals.

Role of ion channels in ionizing radiation-induced cell death.

Neoadjuvant, adjuvant or definitive fractionated radiation therapy are implemented in first line anti-cancer treatment regimens of many tumor entities. Ionizing radiation kills the tumor cells mainly by causing double strand breaks of their DNA through formation of intermediate radicals. Survival of the tumor cells depends on both, their capacity of oxidative defense and their efficacy of DNA repair. By damaging the targeted cells, ionizing radiation triggers a plethora of stress responses. Among those is the modulation of ion channels such as Ca2+-activated K+ channels or Ca2+-permeable nonselective cation channels belonging to the super-family of transient receptor potential channels. Radiogenic activation of these channels may contribute to radiogenic cell death as well as to DNA repair, glucose fueling, radiogenic hypermigration or lowering of the oxidative stress burden. The present review article introduces these channels and summarizes our current knowledge on the mechanisms underlying radiogenic ion channel modulation. This article is part of a Special Issue entitled: Membrane channels and transporters in cancers.

Structural basis of allosteric interactions among Ca2+-binding sites in a K+ channel RCK domain.

Ligand binding sites within proteins can interact by allosteric mechanisms to modulate binding affinities and control protein function. Here we present crystal structures of the regulator of K+ conductance (RCK) domain from a K+ channel, MthK, which reveal the structural basis of allosteric coupling between two Ca2+ regulatory sites within the domain. Comparison of RCK domain crystal structures in a range of conformations and with different numbers of regulatory Ca2+ ions bound, combined with complementary electrophysiological analysis of channel gating, suggests chemical interactions that are important for modulation of ligand binding and subsequent channel opening.

Calcium-activated potassium channel of the tobacco hornworm, Manduca sexta: molecular characterization and expression analysis.

Large-conductance calcium- and voltage-gated potassium channels (BK or Slowpoke) serve as dynamic integrators linking electrical signaling and intracellular activity. These channels can mediate many different Ca2+-dependent physiological processes including the regulation of neuronal and neuroendocrine cell excitability and muscle contraction. To gain insights into the function of BK channels in vivo, we isolated a full-length cDNA encoding the alpha subunit of a Slowpoke channel from the tobacco hornworm, Manduca sexta (msslo). Amino acid sequence comparison of the deduced Manduca protein revealed at least 80% identity to the insect Slo channels. The five C-terminal alternative splice regions are conserved, but the cloned cDNA fragments contained some unique combinations of exons E, G and I. Our spatial profile revealed that transcript levels were highest in skeletal muscle when compared with the central nervous system (CNS) and visceral muscle. The temporal profile suggested that msslo expression is regulated developmentally in a tissue- and regional-specific pattern. The levels of msslo transcripts remain relatively constant throughout metamorphosis in the CNS, transiently decline in the heart and are barely detectable in the gut except in adults. A dramatic upregulation of msslo transcript levels occurs in thoracic but not abdominal dorsal longitudinal body wall muscles (DLM), suggesting that the msSlo current plays an important role in the excitation or contractile properties of the phasic flight muscle. Our developmental profile of msslo expression suggests that msSlo currents may contribute to the changes in neural circuits and muscle properties that produce stage-specific functions and behaviors.

4-Chlorobenzo[F]isoquinoline (CBIQ), a novel activator of CFTR and DeltaF508 CFTR.

4-Chlorobenzo[F]isoquinoline (CBIQ) is a novel compound, here shown to activate both CFTR (cystic fibrosis transmembrane conductance regulator) Cl- ion channels and KCNN4, intermediate conductance, calcium-sensitive K+-channels, present in transporting epithelia by the use of heterologous expression systems. Earlier studies with other benzoquinolines, namely 7,8- and 5,6 benzoquinoline, showed they too could activate CFTR and KCNN4, but the evidence was only indirect. However this study also shows that CBIQ can also activate DeltaF508 CFTR, the most common mutant form of CFTR present in approximately 75% of patients with cystic fibrosis. This property is not shared with the other benzoquinolines. As activation of CFTR and KCNN4 work in unison to promote epithelial chloride secretion, CBIQ is a new chemical scaffold for developing agents that may be useful in cystic fibrosis.

pH dependence of extracellular calcium sensing receptor activity determined by a novel technique.

BACKGROUND: Increasing evidence points to the role of the extracellular Calcium Sensing Receptor (CaSR) as a multimodal receptor responding to diverse physiologic stimuli, such as extracellular divalent and polyvalent cations, amino acids, and ionic strength. Within the kidney, these stimuli converge on the CaSR to coordinate systemic calcium and water homeostasis. In this process, the impact of urinary pH changes on the activity of the CaSR has not yet been defined. We therefore performed the present study to analyze the pH sensitivity of the CaSR. METHODS: To assess the activation state of the CaSR, we developed a new method based on the functional coupling between CaSR activity and gating of calcium sensitive potassium currents mediated by SK4 potassium channels. Two-electrode voltage clamping was used to determine whole cell currents in Xenopus oocytes heterologously expressing rat CaSR and rat SK4 potassium channels. RESULTS: Coexpression of CaSR and SK4 gave rise to potassium currents that were dependent on CaSR-mediated intracellular calcium release, and thereby corresponded to the activation state of the CaSR. In presence of extracellular calcium, ambient alkalinization above pH 7.5 increased CaSR activity. Evaluation of the CaSR calcium sensitivity at various ambient proton concentrations revealed that this effect was due to a sensitization of the CaSR towards extracellular calcium. CONCLUSION: Coexpression with SK4 potassium channels provides a fast and sensitive approach to evaluate CaSR activity in Xenopus oocytes. As disclosed by this novel technique, CaSR activity is regulated by extracellular pH.

Intracorporal injection of hSlo cDNA restores erectile capacity in STZ-diabetic F-344 rats in vivo.

The ability of gene transfer with the pore-forming subunit of the human maxi-K channel (hSlo) to ameliorate the decline in erectile capacity commensurate with 12-24 wk of streptozotocin (STZ)-diabetes was examined in 181 Fischer-344 rats. A 2-mo period of STZ-diabetes was induced before gene transfer, and erectile capacity was evaluated by measuring the intracavernous pressure response (ICP) to cavernous nerve (CN) stimulation (ranging from 0.5 to 10 mA). In the first series of experiments, ANOVA revealed increased CN-stimulated ICP responses at 1 and 2 mo postinjection of 100 microg pcDNA-hSlo compared with control values. A second series of experiments further examined the dose dependence and duration of gene transfer. The ICP response to submaximal (0.5 mA) and maximal (10 mA) nerve stimulation was evaluated 3 or 4 mo postinjection of a single dose of pcDNA-hSlo ranging from 10 to 1,000 microg. ANOVA again revealed that hSlo overexpression was associated with increased CN-stimulated ICP responses compared with responses in corresponding control animals. Histological studies revealed no immune response to the presence of hSlo. PCR analysis documented that expression of both plasmid and transcript were largely confined to the corporal tissue. In the third series of pharmacological experiments, hSlo gene transfer in vivo was associated with iberiotoxin-sensitive relaxation responses to sodium nitroprusside in corporal tissue strips in vitro. The latter data indicate that gene transfer produces functional maxi-K channels that participate in the modulation of corporal smooth muscle cell tone. Taken together, these observations suggest a fundamental diabetes-related change in corporal myocyte maxi-K channel regulation, expression, or function that may be corrected by expression of recombinant hSlo.

The appearance of a protein kinase A-regulated splice isoform of slo is associated with the maturation of neurons that control reproductive behavior.

In response to brief synaptic stimulation that activates protein kinase A (PKA), the bag cell neurons of Aplysia trigger the onset of reproductive behaviors by generating a prolonged afterdischarge. In juvenile animals, such afterdischarges are inhibited by a high density of Ca2+ -activated K+ (BK) channels, encoded by the slo gene. An increase in this current also follows an afterdischarge in mature animals, contributing to a subsequent refractory state that limits reproductive behaviors. Using a bag cell cDNA library, we have isolated two alternative transcripts of the slo gene, differing in the presence (slo-a) or absence (slo-b) of a consensus phosphorylation site for PKA. Expression of either isoform in Chinese hamster ovary cells produced Ca2+ - and voltage-dependent channels with macroscopic and unitary properties matching those in bag cell neurons. The isoforms differed, however, in their response to application of the catalytic subunit of PKA, which reduced the open probability of Slo-a, an effect that was reversed by a PKA inhibitor. In contrast, PKA had no effect on Slo-b. By immunocytochemistry, we determined that the PKA-regulated Slo-a subunit is present in adult, but not juvenile, bag cell neurons. Patch clamp recordings from adult and juvenile bag cell neurons confirmed that PKA decreases BK channel activity only in adults. Our findings suggest that a change in the identity of Slo isoforms expressed during development allows mature neurons to generate afterdischarges that are required for reproduction.

MaxiK channel-triggered negative feedback system is preserved in the urinary bladder smooth muscle from streptozotocin-induced diabetic rats.

MaxiK channel, the large-conductance Ca2+-sensitive K+ channel, facilitates a negative feedback mechanism to oppose excitation and contraction in various types of smooth muscles including urinary bladder smooth muscle (UBSM). In this study, we investigated how the contribution of MaxiK channel to the regulation of basal UBSM mechanical activity is altered in streptozotocin-induced diabetic rats. Although the urinary bladder preparations from both control and diabetic rats were almost quiescent in their basal mechanical activities, they generated spontaneous rhythmic contractions in response to a MaxiK channel blocker, iberiotoxin (IbTx). The effect of IbTx on the mechanical activity was significantly greater in diabetic rat than in control animal. Similarly, the basal mechanical activity was increased with apamin, an inhibitor for some types of small conductance Ca2+-sensitive K+ channels, and this effect was more pronounced for diabetic rat. However, in both control and diabetic animals, IbTx action was stronger than that of apamin. Diabetes also enhanced the responses to BayK 8644, an L-type Ca2+ channel agonist. The extent of this enhancement in diabetic bladder vs. control was, however, almost the same as that attained with IbTx. Expression levels for MaxiK channel as well as apamin-sensitive K+ channels and L-type Ca2+ channel were not altered by diabetes, when determined as their corresponding mRNA levels. These results indicate that diabetes can potentially increase the basal UBSM mechanical activity. However, in diabetic UBSM, the main negative-feedback system triggered by MaxiK channel is still preserved enough to counteract the possible enhancement of this smooth muscle mechanical activity.

Comparative immunohistochemical distribution of three small-conductance Ca2+-activated potassium channel subunits, SK1, SK2, and SK3 in mouse brain.

To investigate the distribution of all three SK channel subunits in the mouse central nervous system, we performed immunohistochemistry using sequence-specific antibodies directed against SK1, SK2, and SK3 proteins. Expression of SK1 and SK2 proteins revealed a partly overlapping distribution pattern restricted to a limited number of brain areas (e.g., neocortex, hippocampal formation). In contrast, SK3 immunoreactivity was rather complementary and predominantly detected in phylogenetically older brain regions like basal ganglia, thalamus, and various brain stem nuclei (e.g., locus coeruleus, tegmental nuclei). At the cellular level, SK1- and SK2-like immunoreactivity was primarily localized to somatic and dendritic structures, whereas the majority of SK3-like immunoreactivity was associated with varicose fibers.

SKCa channels mediate the medium but not the slow calcium-activated afterhyperpolarization in cortical neurons.

Many neurons, including pyramidal cells of the cortex, express a slow afterhyperpolarization (sAHP) that regulates their firing. Although initial findings suggested that the current underlying the sAHP could be carried through SK(Ca) channels, recent work has uncovered anomalies that are not congruent with this idea. Here, we used overexpression and dominant-negative strategies to assess the involvement of SK(Ca) channels in mediating the current underlying the sAHP in pyramidal cells of the cerebral cortex. Pyramidal cells of layer V exhibit robust AHP currents composed of two kinetically and pharmacologically distinguishable currents known as the medium AHP current (I(mAHP)) and the slow AHP current (I(sAHP)). I(mAHP) is blocked by the SK(Ca) channel blockers apamin and bicuculline, whereas I(sAHP) is resistant to these agents but is inhibited by activation of muscarinic receptors. To test for a role for SK(Ca) channels, we overexpressed K(Ca)2.1 (SK1) and K(Ca)2.2 (SK2), the predominant SK(Ca) subunits expressed in the cortex, in pyramidal cells of cultured brain slices. Overexpression of K(Ca)2.1 and K(Ca)2.2 resulted in a fourfold to fivefold increase in the amplitude of I(mAHP) but had no detectable effect on I(sAHP). As an additional test, we examined I(sAHP) in a transgenic mouse expressing a truncated SK(Ca) subunit (SK3-1B) capable of acting as a dominant negative for the entire family of SK(Ca)-IK(Ca) channels. Expression of SK3-1B profoundly inhibited I(mAHP) but again had no discernable effect on I(sAHP). These results are inconsistent with the proposal that SK(Ca) channels mediate I(sAHP) in pyramidal cells and indicate that a different potassium channel mediates this current.

Co-expression of mCysLT1 receptors and IK channels in Xenopus laevis oocytes elicits LTD4-stimulated IK current, independent of an increase in [Ca2+]i.

Addition of LTD4 (10 nM) to Xenopus laevis oocytes expressing the mCysLT1 receptor together with hBK or hIK channels resulted in the activation of both channels secondary to an LTD4-induced increase in [Ca2+]i. In addition, the hIK channel is activated by low concentrations of LTD4 (<0.1 nM), which did not result in any increase in [Ca2+]i. Even though activation of hIK by low concentrations of LTD4 was independent of an increase in [Ca2+]i, a certain "permissive" level of [Ca2+]i was required for its activation, since buffering of intracellular Ca2+ by EGTA completely abolished the response to LTD4. Neither hTBAK1 nor hTASK2 was activated following stimulations with LTD4 (0.1 and 100 nM).

SK3-1C, a dominant-negative suppressor of SKCa and IKCa channels.

Small conductance Ca2+-activated K+ channels, products of the SK1-SK3 genes, regulate membrane excitability both within and outside the nervous system. We report the characterization of a SK3 variant (SK3-1C) that differs from SK3 by utilizing an alternative first exon (exon 1C) in place of exon 1A used by SK3, but is otherwise identical to SK3. Quantitative RT-PCR detected abundant expression of SK3-1C transcripts in human lymphoid tissues, skeletal muscle, trachea, and salivary gland but not the nervous system. SK3-1C did not produce functional channels when expressed alone in mammalian cells, but suppressed SK1, SK2, SK3, and IKCa1 channels, but not BKCa or KV channels. Confocal microscopy revealed that SK3-1C sequestered SK3 protein intracellularly. Dominant-inhibitory activity of SK3-1C was not due to a nonspecific calmodulin sponge effect since overexpression of calmodulin did not reverse SK3-1C-mediated intracellular trapping of SK3 protein, and calmodulin-Ca2+-dependent inactivation of CaV channels was not affected by SK3-1C overexpression. Deletion analysis identified a dominant-inhibitory segment in the SK3-1C C terminus that resembles tetramerization-coiled-coiled domains reported to enhance tetramer stability and selectivity of multimerization of many K+ channels. SK3-1C may therefore suppress calmodulin-gated SKCa/IKCa channels by trapping these channel proteins intracellularly via subunit interactions mediated by the dominant-inhibitory segment and thereby reduce functional channel expression on the cell surface. Such family-wide dominant-negative suppression by SK3-1C provides a powerful mechanism to titrate membrane excitability and is a useful approach to define the functional in vivo role of these channels in diverse tissues by their targeted silencing.

Blockade of the intermediate-conductance calcium-activated potassium channel as a new therapeutic strategy for restenosis.

BACKGROUND: Angioplasty stimulates proliferation and migration of vascular smooth muscle cells (VSMC), leading to neointimal thickening and vascular restenosis. In a rat model of balloon catheter injury (BCI), we investigated whether alterations in expression of Ca2+-activated K+ channels (KCa) contribute to intimal hyperplasia and vascular restenosis. METHODS AND RESULTS: Function and expression of KCa in mature medial and neointimal VSMC were characterized in situ by combined single-cell RT-PCR and patch-clamp analysis. Mature medial VSMC exclusively expressed large-conductance KCa (BKCa) channels. Two weeks after BCI, expression of BKCa was significantly reduced in neointimal VSMC, whereas expression of intermediate-conductance KCa (IKCa1) channels was upregulated. In the aortic VSMC cell line, A7r5 epidermal growth factor (EGF) induced IKCa1 upregulation and EGF-stimulated proliferation was suppressed by the selective IKCa1 blocker TRAM-34. Daily in vivo administration of TRAM-34 to rats significantly reduced intimal hyperplasia by approximately 40% at 1, 2, and 6 weeks after BCI. Two weeks of treatment with the related compound clotrimazole was equally effective. Reduction of intimal hyperplasia was accompanied by decreased neointimal cell content, with no change in the rate of apoptosis or collagen content. CONCLUSIONS: The switch toward IKCa1 expression may promote excessive neointimal VSMC proliferation. Blockade of IKCa1 could therefore represent a new therapeutic strategy to prevent restenosis after angioplasty.

Selective regulation of xSlo splice variants during Xenopus embryogenesis.

Calcium-activated potassium channels regulate excitability of the adult nervous system. In contrast, little is known about the contribution of calcium-activated potassium channels to excitability of the embryonic nervous system when electrical membrane properties and intracellular calcium levels show dramatic changes. Embryonic Xenopus spinal neurons exhibit a well-characterized developmental program of excitability that involves several different currents including calcium-activated ones. Here, we show that a molecular determinant of calcium-activated potassium channels, xSlo, is expressed during Xenopus embryogenesis even prior to differentiation of excitable tissues. Five different xSlo variants are expressed in embryonic tissues as a consequence of alternative exon usage at a single splice site. One of these variants, xSlo59, is neural-specific, and its expression is limited to late stages of neuronal differentiation. However, expression of the four other variants occurs in both muscle and neurons at all stages of development examined. Electrophysiological analysis of recombinant xSlo channels reveals that the xSlo59 exon serves as a gain-of-function module and allows physiologically relevant levels of membrane potential and intracellular calcium to activate effectively the resultant channel. These results suggest that xSlo59 channels play a unique role in sculpting the excitable membrane properties of Xenopus spinal neurons.

The large conductance potassium channel beta-subunit can interact with and modulate the functional properties of a calcium-activated chloride channel, CLCA1.

We have recently compared the biophysical and pharmacological properties of native Ca(2+)-activated Cl(-) currents in murine portal vein with mCLCA1 channels cloned from murine portal vein myocytes (Britton, F. C., Ohya, S., Horowitz, B., and Greenwood, I. A. (2002) J. Physiol. (Lond.) 539, 107-117). These channels shared a similar relative permeability to various anions, but the expressed channel current lacked the marked time dependence of the native current. In addition, the expressed channel showed a lower Ca(2+) sensitivity than the native channel. As non-pore-forming regulatory beta-subunits alter the kinetics and increase the Ca(2+) sensitivity of Ca(2+)-dependent K(+) channels (BK channels) we investigated whether co-expression of beta-subunits with CLCA1 would alter the kinetics/Ca(2+) sensitivity of mCLCA1. Internal dialysis of human embryonic kidney cells stably expressing CLCA1 with 500 nM Ca(2+) evoked a significantly larger current when the beta-subunit KCNMB1 was co-expressed. In a small number of co-transfected cells marked time dependence to the activation kinetics was observed. Interaction studies using the mammalian two-hybrid technique demonstrated a physical association between CLCA1 and KCNMB1 when co-expressed in human embryonic kidney cells. These data suggest that activation of CLCA1 can be modified by accessory subunits.

Steady-state and closed-state inactivation properties of inactivating BK channels.

Calcium-dependent potassium (BK-type) Ca2+ and voltage-dependent K+ channels in chromaffin cells exhibit an inactivation that probably arises from coassembly of Slo1 alpha subunits with auxiliary beta subunits. One goal of this work was to determine whether the Ca2+ dependence of inactivation arises from any mechanism other than coupling of inactivation to the Ca2+ dependence of activation. Steady-state inactivation and the onset of inactivation were studied in inside-out patches and whole-cell recordings from rat adrenal chromaffin cells with parallel experiments on inactivating BK channels resulting from cloned alpha + beta2 subunits. In both cases, steady-state inactivation was shifted to more negative potentials by increases in submembrane [Ca2+] from 1 to 60 microM. At 10 and 60 microM Ca2+, the maximal channel availability at negative potentials was similar despite a shift in the voltage of half availability, suggesting there is no strictly Ca2+-dependent inactivation. In contrast, in the absence of Ca2+, depolarization to potentials positive to +20 mV induces channel inactivation. Thus, voltage-dependent, but not solely Ca2+-dependent, kinetic steps are required for inactivation to occur. Finally, under some conditions, BK channels are shown to inactivate as readily from closed states as from open states, indicative that a key conformational change required for inactivation precedes channel opening.

Cloning and characterization of glioma BK, a novel BK channel isoform highly expressed in human glioma cells.

Voltage-dependent large-conductance Ca2+-activated K+ channels (BK channels) are widely expressed in excitable and nonexcitable cells. BK channels exhibit diverse electrophysiological properties, which are attributable in part to alternative splicing of their alpha-subunits. BK currents have been implicated in the growth control of glial cells, and BK channels with novel biophysical properties have recently been characterized in human glioma cells. Here we report the isolation, cloning, and functional characterization of glioma BK (gBK), a novel splice isoform of hSlo, the gene that encodes the alpha-subunits of human BK channels. The primary sequence of gBK is 97% identical to its closest homolog hbr5, but it contains an additional 34-amino-acid exon at splice site 2 in the C-terminal tail of BK channels. hSlo transcripts containing this novel exon are expressed ubiquitously in various normal tissues as well as in neoplasmic samples, suggesting that the novel exon may modulate important physiological functions of BK channels. Expression of gBK in Xenopus oocytes gives rise to iberiotoxin-sensitive (IbTX) currents, with an IC(50) for IbTX of 5.7 nm and a Hill coefficient of 0.76. Single gBK channels have a unitary conductance of similar250 pS, and the currents show significantly slower activation and higher Ca2+ sensitivity than hbr5. Ca2+ sensitivity was enhanced specifically at physiologically relevant [Ca2+]i (100-500 nm). Examination of biopsies from patients with malignant gliomas has revealed specific overexpression of BK channels in gliomas compared with nonmalignant human cortical tissues. Importantly, tumor malignancy grades have correlated positively with BK channel expression, suggesting an important role for the gBK channel in glioma biology.