5,6-EET potently inhibits T-type calcium channels: implication in the regulation of the vascular tone

T-type calcium channels (T-channels) are important actors in neuronal pacemaking, in heart rhythm, and in the control of the vascular tone. T-channels are regulated by several endogenous lipids including the primary eicosanoid arachidonic acid (AA), which display an important role in vasodilation via its metabolism leading to prostanoids, leukotrienes, and epoxyeicosatrienoic acids (EETs). However, the effects of these latter molecules on T-currents have not been investigated. Here, we describe the effects of the major cyclooxygenase, lipoxygenase, and cytochrome P450 epoxygenase products on the three human recombinant T-channels (Ca_v3.1, Ca_v3.2, and Ca_v3.3), as compared to those of AA. We identified the P450 epoxygenase product, 5,6-EET, as a potent physiological inhibitor of Ca_v3 currents. The effects of 5,6-EET were observed at sub-micromolar concentrations (IC₅₀?=?0.54 μM), occurred in the minute range, and were reversible. The 5,6-EET inhibited the Ca_v3 currents at physiological resting membrane potentials mostly by inducing a large negative shift in their steady-state inactivation properties. Using knockout mice for Ca_v3.1 and Ca_v3.2, we demonstrated that the vasodilation of preconstricted mesenteric arteries induced by 5,6-EET was specifically impaired in Ca_v3.2 knockout mice. Overall, our results indicate that inhibition of Ca_v3 currents by 5,6-EET is an important mechanism controlling the vascular tone.     

CaV3.2 calcium channels control NMDA receptor-mediated transmission: a new mechanism for absence epilepsy

Ca_V3.2 T-type calcium channels, encoded by CACNA1H, are expressed throughout the brain, yet their general function remains unclear. We discovered that Ca_V3.2 channels control NMDA-sensitive glutamatergic receptor (NMDA-R)-mediated transmission and subsequent NMDA-R-dependent plasticity of AMPA-R-mediated transmission at rat central synapses. Interestingly, functional Ca_V3.2 channels primarily incorporate into synapses, replace existing Ca_V3.2 channels, and can induce local calcium influx to control NMDA transmission strength in an activity-dependent manner. Moreover, human childhood absence epilepsy (CAE)-linked hCa_V3.2(C456S) mutant channels have a higher channel open probability, induce more calcium influx, and enhance glutamatergic transmission. Remarkably, cortical expression of hCa_V3.2(C456S) channels in rats induces 2- to 4-Hz spike and wave discharges and absence-like epilepsy characteristic of CAE patients, which can be suppressed by AMPA-R and NMDA-R antagonists but not T-type calcium channel antagonists. These results reveal an unexpected role of Ca_V3.2 channels in regulating NMDA-R-mediated transmission and a novel epileptogenic mechanism for human CAE.     

Regulation of the mitochondrial proton gradient by cytosolic Ca2+ signals

Mitochondria convert the energy stored in carbohydrate and fat into ATP molecules that power enzymatic reactions within cells, and this process influences cellular calcium signals in several ways. By providing ATP to calcium pumps at the plasma and intracellular membranes, mitochondria power the calcium gradients that drive the release of Ca²⁺ from stores and the entry of Ca²⁺ across plasma membrane channels. By taking up and subsequently releasing calcium ions, mitochondria determine the spatiotemporal profile of cellular Ca²⁺ signals and the activity of Ca²⁺-regulated proteins, including Ca²⁺ entry channels that are themselves part of the Ca²⁺ circuitry. Ca²⁺ elevations in the mitochondrial matrix, in turn, activate Ca²⁺-dependent enzymes that boost the respiratory chain, increasing the ability of mitochondria to buffer calcium ions. Mitochondria are able to encode and decode Ca²⁺ signals because the respiratory chain generates an electrochemical gradient for protons across the inner mitochondrial membrane. This proton motive force (??p) drives the activity of the ATP synthase and has both an electrical component, the mitochondrial membrane potential (???? _m ), and a chemical component, the mitochondrial proton gradient (??pH _m ). ???? _m contributes about 190?mV to ??p and drives the entry of Ca²⁺ across a recently identified Ca²⁺-selective channel known as the mitochondrial Ca²⁺ uniporter. ??pH _m contributes ~30?mV to ??p and is usually ignored or considered a minor component of mitochondria respiratory state. However, the mitochondrial proton gradient is an essential component of the chemiosmotic theory formulated by Peter Mitchell in 1961 as ??pH _m sustains the entry of substrates and metabolites required for the activity of the respiratory chain and drives the activity of electroneutral ion exchangers that allow mitochondria to maintain their osmolarity and volume. In this review, we summarize the mechanisms that regulate the mitochondrial proton gradient and discuss how thermodynamic concepts derived from measurements in purified mitochondria can be reconciled with our recent findings that mitochondria have high proton permeability in situ and that ??pH _m decreases during mitochondrial Ca²⁺ elevations.     

Neuropathic pain: role for presynaptic T-type channels in nociceptive signaling

Pain is an important clinical problem and, in its chronic form, may be a disabling condition. Most currently available therapies are insufficient and/or accompanied by serious side effects. Recent studies have implicated the Ca_V3.2 isoform of T-type Ca channels in nociceptive signaling. Ca_V3.2 channels are located in the somas of dorsal root ganglion cells and in the central endings of these cells in the dorsal horn of the spinal cord. These channels can support the development and maintenance of both physiological (nociceptive) and pathological (neuropathic) pain. In this review, we summarize the most recent evidence linking the presynaptic Ca_V3.2 channels to the etiology of neuropathic pain disorders. In particular, we focus on data linking plasticity of Ca_V3.2 channels with neuropathic pain disorders associated with mechanical peripheral nerve injury and with diabetic peripheral neuropathy. We also discuss the development of potential pain therapies aimed at these channels.     

CaV2.1 channels are modulated by muscarinic M1 receptors through phosphoinositide hydrolysis in neostriatal neurons

In adult neostriatal projection neurons, the intracellular Ca²⁺ supplied by Ca_V2.1 (P/Q) Ca²⁺ channels is in charge of both the generation of the afterhyperpolarizing potential (AHP) and the release of GABA from their synaptic terminals, thus being a major target for firing pattern and transmitter release modulations. We have shown that activation of muscarinic M₁-class receptors modulates Ca_V2.1 channels in these neurons in rats. This modulation is reversible, is not membrane delimited, is blocked by the specific M₁-class muscarinic antagonist muscarine toxin 7 (MT-7), and is neither mediated by protein kinase C (PKC) nor by protein phosphatase 2B (PP-2B). Hence, the signaling mechanism of muscarinic Ca_V2.1 channel modulation has remained elusive. The present paper shows that inactivation of phospholipase C (PLC) abolishes this modulation while inhibition of phosphoinositide kinases, PI-3K and PI-4K, prevents its reversibility, suggesting that the reconstitution of muscarinic modulation depends on phosphoinositide rephosphorylation. In support of this hypothesis, the supply of intracellular phosphatidylinositol (4,5) bisphosphate [PI(4,5)P₂] blocked all muscarinic modulation of this channel. The results indicate that muscarinic M₁ modulation of Ca_V2.1 Ca²⁺ channels in these neurons involves phosphoinositide hydrolysis.     

Ca2+ channels,ryanodine receptors and Ca2+-activated K+ channels: a functional unit for regulating arterial tone

Local calcium transients (‘Ca²⁺ sparks’) are thought to be elementary Ca²⁺ signals in heart, skeletal and smooth muscle cells. Ca²⁺ sparks result from the opening of a single, or the coordinated opening of many, tightly clustered ryanodine receptor (RyR) channels in the sarcoplasmic reticulum (SR). In arterial smooth muscle, Ca²⁺ sparks appear to be involved in opposing the tonic contraction of the blood vessel. Intravascular pressure causes a graded membrane potential depolarization to approximately ?40 mV, an elevation of arterial wall [Ca²⁺]_i and contraction (‘myogenic tone’) of arteries. Ca²⁺ sparks activate calcium-sensitive K⁺ (K_Ca) channels in the sarcolemmal membrane to cause membrane hyperpolarization, which opposes the pressure induced depolarization. Thus, inhibition of Ca²⁺ sparks by ryanodine, or of K_Ca channels by iberiotoxin, leads to membrane depolarization, activation of L -type voltage-gated Ca²⁺ channels, and vasoconstriction. Conversely, activation of Ca²⁺ sparks can lead to vasodilation through activation of K_Ca channels. Our recent work is aimed at studying the properties and roles of Ca²⁺ sparks in the regulation of arterial smooth muscle function. The modulation of Ca²⁺ spark frequency and amplitude by membrane potential, cyclic nucleotides and protein kinase C will be explored. The role of local Ca²⁺ entry through voltage-dependent Ca²⁺ channels in the regulation of Ca²⁺ spark properties will also be examined. Finally, using functional evidence from cardiac myocytes, and histological evidence from smooth muscle, we shall explore whether Ca²⁺ channels, RyR channels, and K_Ca channels function as a coupled unit, through Ca²⁺ and voltage, to regulate arterial smooth muscle membrane potential and vascular tone.     

Amitriptyline modulates calcium currents and intracellular calcium concentration in mouse trigeminal ganglion neurons

Migraine is increasingly recognized as a channelopathy, and abnormalities of voltage-activated ionic channels could represent the molecular basis for the altered neuronal functioning. The high-voltage-activated (HVA) Ca²⁺ channels in the trigeminovascular system play a role in the pathophysiology of migraine. In the present study, effects of amitriptyline (AMT), a commonly used migraine prophylactic drug, on the HVA calcium currents (I_Ca) were examined in mouse trigeminal ganglion neurons using whole-cell patch clamp technique. AMT produced concentration- and use-dependent inhibition of HVA I_Ca. Bath application of GÖ-6983 (a selective protein kinase C inhibitor) or H89 (a protein kinase A inhibitor) did not reduce the AMT-induced inhibition of HVA I_Ca. A similar inhibition was observed when calcium imaging was used to directly monitor the effects of AMT on KCl-induced increments of intracellular Ca²⁺ concentration ([Ca²⁺]_i). By blocking HVA Ca²⁺ channels and Ca²⁺ entry into cells, AMT could prevent the release of neurotransmitters and help restore the neuronal threshold for excitation. Our findings suggest interesting therapeutic mechanisms for AMT in migraine prevention.     

Cysteines in the loop between IS5 and the pore helix of CaV3.1 are essential for channel gating

The role of six cysteines of Ca_V3.1 in channel gating was investigated. C241, C271, C282, C298, C313, and C323, located in the extracellular loop between segment IS5 and the pore helix, were each mutated to alanine; the resultant channels were expressed and studied by patch clamping in HEK293 cells. C298A and C313A conducted calcium currents, while the other mutants were not functional. C298A and C313A as well as double mutation C298/313A significantly reduced the amplitude of the calcium currents, shifted the activation curve in the depolarizing direction and slowed down channel inactivation. Redox agents dithiothreitol (DTT) and 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) shifted the current activation curve of wild-type channels in the hyperpolarizing direction. Activation curve for all mutated channels was shifted in hyperpolarizing direction by DTT while DTNB caused a depolarizing shift. Our study reveals that the cysteines we studied have an essential role in Ca_V3.1 gating. We hypothesize that cysteines in the large extracellular loop of Ca_V3.1 form bridges within the loop and/or neighboring channel segments that are essential for channel gating.     

Properties and role of voltage-dependent calcium channels during mouse skeletal muscle differentiation

Skeletal muscle differentiation depends on calcium ions, but it is yet unclear whether calcium entry through voltage-dependent calcium channels (VDCCs) contributes to the myoblast fusion process. In this study, we investigate whether calcium influx through functional T-type VDCCs precedes and affects mouse satellite cell fusion. We report here on the properties and the role of the VDCCs expressed in differentiating mouse muscular cells using both the C2C12 cell line and primary cultures of satellite cells. We present electrophysiological and biochemical evidence demonstrating that T-type and L-type VDCCs are not present in C2C12 and primary cultures of mouse satellite cells prior to the fusion stage. Although mRNA for the T-type Ca_V3.2 subunit was detected in differentiated C2C12 cells, no T-type calcium currents could be recorded, while both T-type and L-type calcium currents were detected after the fusion process in primary cultures. In addition, chronic application of 30 μM nickel, known to inhibit T-type Ca_V3.2 channels, did not alter the fusion of C2C12 cells and mouse satellite cells in primary culture. Overall, the data indicate that, unlike in humans, Ca_V3.2 T-type calcium channels play no role in mouse satellite cell fusion.     

K+ channel modulation in arterial smooth muscle

Potassium channels play an essential role in the membrane potential of arterial smooth muscle, and also in regulating contractile tone. Four types of K⁺ channel have been described in vascular smooth muscle: Voltage-activated K⁺ channels (K_V) are encoded by the Kv gene family, Ca²⁺-activated K⁺ channels (BK_Ca) are encoded by the slogene, inward rectifiers (K_IR) by Kir2.0, and ATP-sensitive K⁺ channels (K_ATP) by Kir6.0 and sulphonylurea receptor genes. In smooth muscle, the channel subunit genes reported to be expressed are: Kv1.0, Kv1.2, Kv1.4–1.6, Kv2.1, Kv9.3, Kvβ1–β4, slo α and β, Kir2.1, Kir6.2, and SUR1 and SUR2. Arterial K⁺ channels are modulated by physiological vasodilators, which increase K⁺ channel activity, and vasoconstrictors, which decrease it. Several vasodilators acting at receptors linked to cAMP-dependent protein kinase activate K_ATP channels. These include adenosine, calcitonin gene-related peptide, and β-adrenoceptor agonists. β-adrenoceptors can also activate BK_Ca and K_V channels. Several vasoconstrictors that activate protein kinase C inhibit K_ATP channels, and inhibition of BK_Ca and K_V channels through PKC has also been described. Activators of cGMP-dependent protein kinase, in particular NO, activate BK_Ca channels, and possibly K_ATP channels. Hypoxia leads to activation of K_ATP channels, and activation of BK_Ca channels has also been reported. Hypoxic pulmonary vasoconstriction involves inhibition of K_V channels. Vasodilation to increased external K⁺ involves K_IR channels. Endothelium-derived hyperpolarizing factor activates K⁺ channels that are not yet clearly defined. Such K⁺ channel modulations, through their effects on membrane potential and contractile tone, make important contributions to the regulation of blood flow.     

T-type Ca2+ channels in spermatogenic cells and sperm

Cell function is importantly regulated by the intracellular concentration of Ca²⁺ ([Ca²⁺]i). Sperm development and function are deeply influenced by [Ca²⁺]i which is modulated amongst other ion transporters by plasma membrane Ca²⁺ permeable channels. The presence and role of voltage-dependent Ca²⁺ channels (Ca_V) of the T-type (Ca_V3) in sperm physiology have become a matter of debate in recent years. Though they are functionally present in later stages of development in spermatogenic cells and testicular sperm and their mRNAs and proteins detected from spermatogenic cells to mature mammalian spermatozoa, their currents have not been recorded in mature spermatozoa. This review critically summarizes the evidence for the involvement of Ca_V3 channels in sperm development and function.     

BK<Subscript>Ca</Subscript> and K<Subscript>V</Subscript> channels limit conducted vasomotor responses in rat mesenteric terminal arterioles

Intracellular Ca²⁺ signals underlying conducted vasoconstriction to local application of a brief depolarizing KCl stimulus was investigated in rat mesenteric terminal arterioles (<40 μm). Using a computer model of an arteriole segment comprised of coupled endothelial cells (EC) and vascular smooth muscle cells (VSMC) simulations of both membrane potential and intracellular [Ca²⁺] were performed. The “characteristic” length constant, λ, was approximated using a modified cable equation in both experiments and simulations. We hypothesized that K⁺ conductance in the arteriolar wall limit the electrotonic spread of a local depolarization along arterioles by current dissipation across the VSMC plasma membrane. Thus, we anticipated an increased λ by inhibition of voltage-activated K⁺ channels. Application of the BK_Ca channel blocker iberiotoxin (100 nM) onto mesenteric arterioles in vitro and inhibition of BK_Ca channel current in silico increased λ by 34% and 32%, respectively. Similarly, inhibition of K_V channels in vitro (4-aminopyridine, 1 mM) or in silico increased λ by 41% and 21%, respectively. Immunofluorescence microscopy demonstrated expression of BK_Ca, Kv1.5, Kv2.1, but not Kv1.2, in VSMCs of rat mesenteric terminal arterioles. Our results demonstrate that inhibition of voltage-activated K⁺ channels enhance vascular-conducted responses to local depolarization in terminal arterioles by increasing the membrane resistance of VSMCs. These data contribute to our understanding of how differential expression patterns of voltage-activated K⁺ channels may influence conducted vasoconstriction in small arteriolar networks. This finding is potentially relevant to understanding the compromised microcirculatory blood flow in systemic vascular diseases such as diabetes mellitus and hypertension.     

Cl− channels in smooth muscle cells

In smooth muscle cells (SMCs), the intracellular chloride ion (Cl^?) concentration is high due to accumulation by Cl^?/HCO₃ ^? exchange and Na⁺–K⁺–Cl^? cotransportation. The equilibrium potential for Cl^? (E _Cl) is more positive than physiological membrane potentials (E _m), with Cl^? efflux inducing membrane depolarization. Early studies used electrophysiology and nonspecific antagonists to study the physiological relevance of Cl^? channels in SMCs. More recent reports have incorporated molecular biological approaches to identify and determine the functional significance of several different Cl^? channels. Both “classic” and cGMP-dependent calcium (Ca²⁺)-activated (Cl_Ca) channels and volume-sensitive Cl^? channels are present, with TMEM16A/ANO1, bestrophins, and ClC-3, respectively, proposed as molecular candidates for these channels. The cystic fibrosis transmembrane conductance regulator (CFTR) has also been described in SMCs. This review will focus on discussing recent progress made in identifying each of these Cl^? channels in SMCs, their physiological functions, and contribution to diseases that modify contraction, apoptosis, and cell proliferation.     

BK channels in intact clonal rat pituitary cells are activated by physiological elevations of the cytosolic Ca2+ concentration at the normal resting potential

Activation of large conductance Ca²⁺-activated K⁺ channels (BK channels) in intact clonal rat pituitary cells (GH₄ cells) was investigated using the cell-attached patch-clamp configuration. This method prevents loss of intracellular factors which might influence channel activity. BK channels are generally considered to be inactive at the resting membrane potential in excitable cells. However, at the resting potential (0 mV pipette potential), 40% of the cell-attached patches displayed spontaneously active BK channels, which remained active even at 20 mV hyperpolarization. The peptide thyroliberin (TRH) elevates the cytosolic Ca²⁺ concentration ([ Ca²⁺]_i) in GH cells by IP₃-induced release of Ca²⁺ from intracellular stores. This rise in [Ca²⁺]_i occurs concomitantly with membrane hyperpolarization. TRH stimulation caused activation of BK channels in nine out of 30 silent cell-attached patches, and caused enhanced channel activity in seven out of 29 cell-attached patches containing spontaneously active BK channels. The Ca²⁺ ionophore ionomycin activated silent BK channels in three out of 10 cell-attached patches, and increased the activity of spontaneously active BK channels in seven out of 16 cell-attached patches. The pipette potential was clamped to 0 mV in all these experiments. We conclude that the BK channels in GH₄ cells may be active at the resting membrane potential and more negative membrane potentials. The channels may also be activated further by physiological elevations of [Ca²⁺]_i in the same potential range. Our results point towards new possible physiological roles for the BK channels in GH₄ cells. This is in agreement with the emerging picture of BK channels highly sensitive to [Ca²⁺]_i in a wide variety of cell types.     

Voltage-activated ion channels and Ca2+-induced Ca2+ release shape Ca2+ signaling in Merkel cells

Ca²⁺ signaling and neurotransmission modulate touch-evoked responses in Merkel cell–neurite complexes. To identify mechanisms governing these processes, we analyzed voltage-activated ion channels and Ca²⁺ signaling in purified Merkel cells. Merkel cells in the intact skin were specifically labeled by antibodies against voltage-activated Ca²⁺ channels (Ca_V2.1) and voltage- and Ca²⁺-activated K⁺ (BK_Ca) channels. Voltage-clamp recordings revealed small Ca²⁺ currents, which produced Ca²⁺ transients that were amplified sevenfold by Ca²⁺-induced Ca²⁺ release. Merkel cells’ voltage-activated K⁺ currents were carried predominantly by BK_Ca channels with inactivating and non-inactivating components. Thus, Merkel cells, like hair cells, have functionally diverse BK_Ca channels. Finally, blocking K⁺ channels increased response magnitude and dramatically shortened Ca²⁺ transients evoked by mechanical stimulation. Together, these results demonstrate that Ca²⁺ signaling in Merkel cells is governed by the interplay of plasma membrane Ca²⁺ channels, store release and K⁺ channels, and they identify specific signaling mechanisms that may control touch sensitivity.     

Temporal profile of potassium channel dysfunction in cerebrovascular smooth muscle after experimental subarachnoid haemorrhage

The pathogenesis of cerebral vasospasm after subarachnoid haemorrhage (SAH) involves sustained contraction of arterial smooth muscle cells that is maximal 6–8 days after SAH. We reported that function of voltage-gated K⁺ (K_V) channels was significantly decreased during vasospasm 7 days after SAH in dogs. Since arterial constriction is regulated by membrane potential that in turn is determined predominately by K⁺ conductance, the compromised K⁺ channel dysfunction may cause vasospasm. Additional support for this hypothesis would be demonstration that K⁺ channel dysfunction is temporally coincident with vasospasm. To test this hypothesis, SAH was created using the double haemorrhage model in dogs and smooth muscle cells from the basilar artery, which develops vasospasm, were isolated 4 days (early vasospasm), 7 days (during vasospasm) and 21 days (after vasospasm) after SAH and studied using patch-clamp electrophysiology. We investigated the two main K⁺ channels (K_V and large-conductance voltage/Ca²⁺-activated (K_Ca) channels). Electrophysiologic function of K_Ca channels was preserved at all times after SAH. In contrast, function of K_V channels was significantly decreased at all times after SAH. The decrease in cell size and degree of K_V channel dysfunction was maximal 7 days after SAH. The results suggest that K_V channel dysfunction either only partially contributes to vasospasm after SAH or that compensatory mechanisms develop that lead to resolution of vasospasm before K_V channels recover their function.     

Novel vistas of calcium-mediated signalling in the thalamus

Traditionally, the role of calcium ions (Ca²⁺) in thalamic neurons has been viewed as that of electrical charge carriers. Recent experimental findings in thalamic cells have only begun to unravel a highly complex Ca²⁺ signalling network that exploits extra- and intracellular Ca²⁺ sources. In thalamocortical relay neurons, interactions between T-type Ca²⁺ channel activation, Ca²⁺-dependent regulation of adenylyl cyclase activity and the hyperpolarization-activated cation current (I_h) regulate oscillatory burst firing during periods of sleep and generalized epilepsy, while a functional triad between Ca²⁺ influx through high-voltage-activated (most likely L-type) Ca²⁺ channels, Ca²⁺-induced Ca²⁺ release via ryanodine receptors (RyRs) and a repolarizing mechanism (possibly via K⁺ channels of the BK_Ca type) supports tonic spike firing as required during wakefulness. The mechanisms seem to be located mostly at dendritic and somatic sites, respectively. One functional compartment involving local GABAergic interneurons in certain thalamic relay nuclei is the glomerulus, in which the dendritic release of GABA is regulated by Ca²⁺ influx via canonical transient receptor potential channels (TRPC), thereby presumably enabling transmitters of extrathalamic input systems that are coupled to phospholipase C (PLC)-activating receptors to control feed-forward inhibition in the thalamus. Functional interplay between T-type Ca²⁺ channels in dendrites and the A-type K⁺ current controls burst firing, contributing to the range of oscillatory activity observed in these interneurons. GABAergic neurons in the reticular thalamic (RT) nucleus recruit a specific set of Ca²⁺-dependent mechanisms for the generation of rhythmic burst firing, of which a particular T-type Ca²⁺ channel in the dendritic membrane, the Ca²⁺-dependent activation of non-specific cation channels (I_CAN) and of K⁺ channels (SK_Ca type) are key players. Glial Ca²⁺ signalling in the thalamus appears to be a basic mechanism of the dynamic and integrated exchange of information between glial cells and neurons. The conclusion from these observations is that a localized calcium signalling network exists in all neuronal and probably also glial cell types in the thalamus and that this network is dedicated to the precise regulation of the functional mode of the thalamus during various behavioural states.     

CaV2.1 channelopathies

Mutations in the CACNA1A gene that encodes the pore-forming α₁ subunit of human voltage-gated Ca_V2.1 (P/Q-type) Ca²⁺ channels cause several autosomal-dominant neurologic disorders, including familial hemiplegic migraine type 1 (FHM1), episodic ataxia type 2, and spinocerebellar ataxia type 6 (SCA6). For each channelopathy, the review describes the disease phenotype as well as the functional consequences of the disease-causing mutations on recombinant human Ca_V2.1 channels and, in the case of FHM1 and SCA6, on neuronal Ca_V2.1 channels expressed at the endogenous physiological level in knockin mouse models. The effects of FHM1 mutations on cortical spreading depression, the phenomenon underlying migraine aura, and on cortical excitatory and inhibitory synaptic transmission in FHM1 knockin mice are also described, and their implications for the disease mechanism discussed. Moreover, the review describes different ataxic spontaneous cacna1a mouse mutants and the important insights into the cerebellar mechanisms underlying motor dysfunction caused by mutant Ca_V2.1 channels that were obtained from their functional characterization.     

Single-nucleotide polymorphisms in vascular Ca2+-activated K+-channel genes and cardiovascular disease

In the cardiovascular system, Ca²⁺-activated K⁺-channels (KCa) are considered crucial mediators in the control of vascular tone and blood pressure by modulating the membrane potential and shaping Ca²⁺-dependent contraction. Vascular smooth muscle cells express the BK_Ca channel which fine-tunes contractility by providing a negative feedback on Ca²⁺-elevations. BK_Ca channel's ion-conducting α-subunit is encoded by the KCa1.1 gene, and the accessory and Ca²⁺-sensitivity modulating β1-subunit is encoded by the KCNMB1 gene. Vascular endothelial cells express the calmodulin-gated KCa channels IK_Ca (encoded by the KCa3.1 gene) and SK_Ca (encoded by the KCa2.3 gene). These two channels mediate endothelial hyperpolarization and initiate the endothelium-derived hyperpolarizing factor-dilator response. Considering these essential roles of KCa in arterial function, mutations in KCa genes have been suspected to contribute to cardiovascular disease in humans. So far, DNA sequence analysis in the population and patient cohorts has identified single-nucleotide polymorphisms (SNPs) in the BK_Ca β1-subunit gene as well as in the α-subunit gene (KCa1.1). Some of these SNPs produce amino acid exchanges and evoke alterations of channel functions (“gain-of-function” as well as “loss-of-function”). Moreover, the epidemiological studies showed that the presence of the E65K polymorphism in, e.g., BK_Ca β1-subunit gene (producing a “gain-of-function”) lowers the prevalence for severe hypertension and myocardial infarction. Other SNPs in the BK_Ca α-subunit gene and also in the KCa3.1 gene expressed in the endothelium have been suggested to increase the risk of cardiovascular disease. These findings from sequence analysis of human KCa genes, and epidemiological studies thus provide evidence that genetic variations and mutations in KCa channel genes contribute to human cardiovascular disease.