Phylogenetic relationships and further unknown diversity of diplostomids (Diplostomida: Diplostomidae) parasitic in kingfishers.

Kingfishers (Alcedinidae Rafinesque) are common inhabitants of wetlands and are known to be definitive hosts to a wide range of digeneans that parasitize fish as second intermediate hosts. Among these digeneans, members of the Diplostomidae Poirier, 1886 (diplostomids) are particularly common. Recent studies of diplostomids collected from kingfishers have revealed that they are probably more diverse than currently known. This particularly concerns the genera Crassiphiala Van Haitsma, 1925 and Uvulifer Yamaguti, 1934. In the present work, we studied seven diplostomid taxa from kingfishers in Brazil, the USA and the Philippines. Partial DNA sequences of the nuclear large ribosomal subunit (28S) and mitochondrial cytochrome c oxidase I (cox1) genes were obtained, and 28S sequences were used to study the phylogenetic interrelationships of these diplostomids. We provide the first DNA sequences from Uvulifer semicircumcisus Dubois et Rausch, 1950 and a member of Subuvulifer Dubois, 1952. Pseudocrassiphiala n. gen. is erected for a previously recognized species-level lineage of Crassiphiala and a new generic diagnosis of Crassiphiala is provided. Crassiphiala jeffreybelli n. sp., Crassiphiala wecksteini n. sp. and Pseudocrassiphiala tulipifera n. sp. are described, and a description of newly collected, high-quality specimens of Crassiphiala bulboglossa Van Haitsma, 1925 (the type-species of the genus) is provided.

Polycomb-dependent repression of the potassium channel-encoding gene KCNA5 promotes cancer cell survival under conditions of stress.

Relapse after clinical remission remains a leading cause of cancer-associated death. Although the mechanisms of tumor relapse are complex, the ability of cancer cells to survive physiological stress is a prerequisite for recurrence. Ewing sarcoma (ES) and neuroblastoma (NB) are aggressive cancers that frequently relapse after initial remission. In addition, both tumors overexpress the polycomb group (PcG) proteins BMI-1 and EZH2, which contribute to tumorigenicity. We have discovered that ES and NB resist hypoxic stress-induced death and that survival depends on PcG function. Epigenetic repression of developmental programs is the most well-established cancer-associated function of PcG proteins. However, we noted that voltage-gated potassium (Kv) channel genes are also targets of PcG regulation in stem cells. Given the role of potassium in regulating apoptosis, we reasoned that repression of Kv channel genes might have a role in cancer cell survival. Here we describe our novel finding that PcG-dependent repression of the Kv1.5 channel gene KCNA5 contributes to cancer cell survival under conditions of stress. We show that survival of cancer cells in stress is dependent upon suppression of Kv1.5 channel function. The KCNA5 promoter is marked in cancer cells with PcG-dependent chromatin repressive modifications that increase in hypoxia. Genetic and pharmacological inhibition of BMI-1 and EZH2, respectively, restore KCNA5 expression, which sensitizes cells to stress-induced death. In addition, ectopic expression of the Kv1.5 channel induces apoptotic cell death under conditions of hypoxia. These findings identify a novel role for PcG proteins in promoting cancer cell survival via repression of KCNA5.

Defects in neural stem cell proliferation and olfaction in Chd7 deficient mice indicate a mechanism for hyposmia in human CHARGE syndrome.

Mutations in CHD7, a chromodomain gene, are present in a majority of individuals with CHARGE syndrome, a multiple anomaly disorder characterized by ocular Coloboma, Heart defects, Atresia of the choanae, Retarded growth and development, Genital hypoplasia and Ear anomalies. The clinical features of CHARGE syndrome are highly variable and incompletely penetrant. Olfactory dysfunction is a common feature in CHARGE syndrome and has been potentially linked to primary olfactory bulb defects, but no data confirming this mechanistic link have been reported. On the basis of these observations, we hypothesized that loss of Chd7 disrupts mammalian olfactory tissue development and function. We found severe defects in olfaction in individuals with CHD7 mutations and CHARGE, and loss of odor evoked electro-olfactogram responses in Chd7 deficient mice, suggesting reduced olfaction is due to a dysfunctional olfactory epithelium. Chd7 expression was high in basal olfactory epithelial neural stem cells and down-regulated in mature olfactory sensory neurons. We observed smaller olfactory bulbs, reduced olfactory sensory neurons, and disorganized epithelial ultrastructure in Chd7 mutant mice, despite apparently normal functional cilia and sustentacular cells. Significant reductions in the proliferation of neural stem cells and regeneration of olfactory sensory neurons in the mature Chd7(Gt/+) olfactory epithelium indicate critical roles for Chd7 in regulating neurogenesis. These studies provide evidence that mammalian olfactory dysfunction due to Chd7 haploinsufficiency is linked to primary defects in olfactory neural stem cell proliferation and may influence olfactory bulb development.

Molecular basis of hypoxia-induced pulmonary vasoconstriction: role of voltage-gated K+ channels.

The hypoxia-induced membrane depolarization and subsequent constriction of small resistance pulmonary arteries occurs, in part, via inhibition of vascular smooth muscle cell voltage-gated K+ (KV) channels open at the resting membrane potential. Pulmonary arterial smooth muscle cell KV channel expression, antibody-based dissection of the pulmonary arterial smooth muscle cell K+ current, and the O2 sensitivity of cloned KV channels expressed in heterologous expression systems have all been examined to identify the molecular components of the pulmonary arterial O2-sensitive KV current. Likely components include Kv2.1/Kv9.3 and Kv1.2/Kv1.5 heteromeric channels and the Kv3.1b alpha-subunit. Although the mechanism of KV channel inhibition by hypoxia is unknown, it appears that KV alpha-subunits do not sense O2 directly. Rather, they are most likely inhibited through interaction with an unidentified O2 sensor and/or beta-subunit. This review summarizes the role of KV channels in hypoxic pulmonary vasoconstriction, the recent progress toward the identification of KV channel subunits involved in this response, and the possible mechanisms of KV channel regulation by hypoxia.

Isoform-specific localization of voltage-gated K+ channels to distinct lipid raft populations. Targeting of Kv1.5 to caveolae.

The precise subcellular localization of ion channels is often necessary to ensure rapid and efficient integration of both intracellular and extracellular signaling events. Recently, we have identified lipid raft association as a novel mechanism for the subcellular sorting of specific voltage-gated K(+) channels to regions of the membrane rich in signaling complexes. Here, we demonstrate isoform-specific targeting of voltage-gated K(+) (Kv) channels to distinct lipid raft populations with the finding that Kv1.5 specifically targets to caveolae. Multiple lines of evidence indicate that Kv1.5 and Kv2.1 exist in distinct raft domains: 1) channel/raft association shows differential sensitivity to increasing concentrations of Triton X-100; 2) unlike Kv2.1, Kv1.5 colocalizes with caveolin on the cell surface and redistributes with caveolin following microtubule disruption; and 3) immunoisolation of caveolae copurifies Kv1.5 channel. Both depletion of cellular cholesterol and inhibition of sphingolipid synthesis alter Kv1.5 channel function by inducing a hyperpolarizing shift in the voltage dependence of activation and inactivation. The differential targeting of Kv channel subtypes to caveolar and noncaveolar rafts within a single membrane represents a unique mechanism of compartmentalization, which may permit isoform-specific modulation of K(+) channel function.

Differential targeting of Shaker-like potassium channels to lipid rafts.

Ion channel targeting within neuronal and muscle membranes is an important determinant of electrical excitability. Recent evidence suggests that there exists within the membrane specialized microdomains commonly referred to as lipid rafts. These domains are enriched in cholesterol and sphingolipids and concentrate a number of signal transduction proteins such as nitric-oxide synthase, ligand-gated receptors, and multiple protein kinases. Here, we demonstrate that the voltage-gated K(+) channel Kv2.1, but not Kv4.2, targets to lipid rafts in both heterologous expression systems and rat brain. The Kv2.1 association with lipid rafts does not appear to involve caveolin. Depletion of cellular cholesterol alters the buoyancy of the Kv2.1 associated rafts and shifts the midpoint of Kv2.1 inactivation by nearly 40 mV without affecting peak current density or channel activation. The differential targeting of Kv channels to lipid rafts represents a novel mechanism both for the subcellular sorting of K(+) channels to regions of the membrane rich in signaling complexes and for modulating channel properties via alterations in lipid content.

Oxygen sensitivity of cloned voltage-gated K(+) channels expressed in the pulmonary vasculature.

Hypoxic pulmonary vasoconstriction is initiated by inhibiting one or more voltage-gated potassium (Kv) channel in the vascular smooth muscle cells (VSMCs) of the small pulmonary resistance vessels. Although progress has been made in identifying which Kv channel proteins are expressed in pulmonary arterial (PA) VSMCs, there are conflicting reports regarding which channels contribute to the native O(2)-sensitive K(+) current. In this study, we examined the effects of hypoxia on the Kv1.2, Kv1.5, Kv2.1, and Kv9.3 alpha subunits expressed in mouse L cells using the whole-cell patch-clamp technique. Hypoxia (PO(2)= approximately 30 mm Hg) reversibly inhibited Kv1.2 and Kv2.1 currents only at potentials more positive than 30 mV. In contrast, hypoxia did not alter Kv1.5 current. Currents generated by coexpression of Kv2.1 with Kv9.3 alpha subunits were reversibly inhibited by hypoxia in the voltage range of the resting membrane potential (E(M)) of PA VSMCs ( approximately 28% at -40 mV). Coexpression of Kv1.2 and Kv1.5 alpha subunits produced currents that displayed kinetic and pharmacological properties distinct from Kv1.2 and Kv1.5 channels expressed alone. Moreover, hypoxia reversibly inhibited Kv1.2/Kv1.5 current activated at physiologically relevant membrane potentials ( approximately 65% at -40 mV). These results indicate that (1) hypoxia reversibly inhibits Kv1.2 and Kv2.1 but not Kv1.5 homomeric channels, (2) Kv1.2 and 1.5 alpha subunits can assemble to form an O(2)-sensitive heteromeric channel, and (3) only Kv1.2/Kv1.5 and Kv2.1/Kv9.3 heteromeric channels are inhibited by hypoxia in the voltage range of the PA VSMC E(M). Thus, these heteromeric channels are strong candidates for the K(+) channel isoforms initiating hypoxic pulmonary vasoconstriction.

Modulation of Kv channel alpha/beta subunit interactions.

Voltage-gated K(+) channels comprise the largest and most diverse class of ion channels. These channels establish the resting membrane potential and modulate the frequency and duration of action potentials in nerve and muscle, as well as being the targets of several antiarrhythmic drugs in the heart. The multiplicity of Kv channel function is further enhanced through modulation by accessory beta subunits, which confer rapid inactivation, alter current amplitudes, and promote cell surface expression. In addition, alpha/beta interactions are also influenced by second messenger pathways. Recent evidence demonstrates that phosphorylation of Kv channel alpha and/or beta subunits may dramatically affect channel properties. The functional response of different K(+) channel subunits to activation of protein kinases represents not only a means to modulate subunit interactions, but also another mechanism for K(+) channel diversity in vivo.

Angiotensin IV receptor-mediated activation of lung endothelial NOS is associated with vasorelaxation.

The hexapeptide angiotensin (ANG) IV, a metabolic product of ANG II, has been reported to play a functional role in the regulation of blood flow in extrapulmonary tissues. Here, we demonstrate that ANG IV-specific (AT4) receptors are present in porcine pulmonary arterial endothelial cells (PAECs) and that the binding of ANG IV to AT4 receptors can be blocked by its antagonist divalinal ANG IV but not by the ANG II-, AT1-, and AT2-receptor blockers [Sar1,Ile8]ANG II, losartan, and PD-123177, respectively. ANG IV significantly increased endothelial cell constitutive nitric oxide synthase (ecNOS) activity (P < 0.05) as well as cellular cGMP content (P < 0. 001). Western blot analysis revealed that ecNOS protein expression was comparable in control and ANG IV-stimulated cells. Divalinal ANG IV but not [Sar1,Ile8]ANG II, losartan, or PD-123177 inhibited the ANG II- and ANG IV-stimulated increases in ecNOS activity and cGMP content in PAECs. Incubation in the presence of N-nitro-L-arginine methyl ester (L-NAME) or methylene blue but not of indomethacin significantly diminished ANG IV-stimulated as well as basal levels of cGMP (P < 0.001). Similarly, in situ studies with precontracted porcine pulmonary arterial rings showed that ANG IV caused an endothelium-dependent relaxation that was blocked by L-NAME or methylene blue. Collectively, these results demonstrate that ANG IV binds to AT4 receptors, activates ecNOS by posttranscriptional modulation, stimulates cGMP accumulation in PAECs, and causes pulmonary arterial vasodilation, suggesting that ANG IV plays a role in the regulation of blood flow in the pulmonary circulation.

Ion channels in vascular smooth muscle: alterations in essential hypertension.

Essential hypertension is characterized by a near normal cardiac output but an increase in total peripheral resistance. In turn, total peripheral resistance is controlled directly by the diameter of the small arteries and arterioles like those in the kidney. The dynamic regulation of renal vessel diameter is governed by the contractile state of the vascular smooth muscle cells that line the vessel walls. This review addresses the role of a number of different ion channels to initiate and maintain the contractile state of the vascular smooth muscle cells in hypertension and the potential prevention of hypertension through gene therapy. These specific channels include Ca2+, K(Ca), Kv, and Cl- channels. In hypertension, it has been reported that increased activity of Ca2+ channels and decreased activity of Kv channels are responsible for the increased contractile tone and resting membrane potential observed in dissociated vascular smooth muscle cells from the spontaneously hypertensive rat. In contrast, increased activity of K(Ca) channels in vascular smooth muscle cells of the SHR has been hypothesized to dampen or brake the activity of Ca2+ and Kv channels. Finally, recent evidence suggests that introducing angiotensin II type-1 receptor antisense into prehypertensive rat pups prevents the onset of pathophysiological alterations observed in hypertension including K+ channel alterations. These results suggest that gene therapy may be a useful pharmacological and physiological tool to combat hypertension.

Prevention of renovascular and cardiac pathophysiological changes in hypertension by angiotensin II type 1 receptor antisense gene therapy.

Hypertension produces pathophysiological changes that are often responsible for the mortality associated with the disease. However, it is unclear whether normalizing blood pressure (BP) with conventional therapy is effective in reversing the pathophysiological damage. The duration and initiation of treatment, site of administration, and agent used all appear to influence the reversal of the pathophysiological alterations associated with hypertension. We have previously established that retrovirally mediated delivery of angiotensin II type 1 receptor antisense (AT1R-AS) attenuates the development of high BP in the spontaneously hypertensive (SH) rat model of human essential hypertension. Our objective was to determine whether this attenuation of high BP is associated with prevention of other pathophysiological changes induced by the hypertensive state. Intracardiac delivery of AT1R-AS in neonates prevented the development of hypertension in SH rats for at least 120 days. Contractile experiments demonstrated an impaired endothelium-dependent vascular relaxation (acetylcholine) and an enhanced contractile response to vasoactive agents (phenylephrine and KCl) in the SH rat renal vasculature. In addition, the voltage-dependent K+ current density, which is believed to contribute to smooth muscle resting membrane potential and basal tone, was decreased in renal resistance artery cells of the SH rat. AT1R-AS treatment prevented each of these renal vascular alterations. Finally, AT1R-AS delivery prevented the pathological alterations observed in the SH rat myocardium, including left ventricular hypertrophy, multifocal fibrosis, and perivascular fibrosis. These observations demonstrate that viral-mediated delivery of AT1R-AS attenuates the development of hypertension on a long term basis, and this is associated with prevention of pathophysiological changes in SH rats.

Modulation of atrioventricular nodal function by metabolic and allosteric regulators of endogenous adenosine in guinea pig heart.

BACKGROUND: There has been increasing interest in the development of agents that utilize endogenous adenosine to exert their actions. We tested the hypothesis that substances that either potentiate the activity (allosteric enhancers) or increase the interstitial concentration (inhibitors of metabolism) of endogenous adenosine may cause event (tachycardia)-specific depression of AV nodal conduction. METHODS AND RESULTS: The frequency-dependent effects of iodotubercidin (ITU, an inhibitor of adenosine kinase), erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA, an inhibitor of adenosine deaminase), draflazine (a nucleoside transport blocker), and PD81,723 (an allosteric enhancer of the A1 adenosine receptor binding) on the stimulus-to-His bundle (SH) interval, a measure of AV nodal conduction, were determined in guinea pig hearts and compared with those of adenosine and diltiazem. All drugs depressed AV nodal conduction in a frequency-dependent manner. The ratios of SH interval prolongations at fast to slow pacing rates for draflazine, ITU + EHNA, PD81,723, adenosine, and diltiazem were 17.5 +/- 3.4, 11.1 +/- 5.0, 3.5 +/- 0.9, 10.1 +/- 2.8, and 8.3 +/- 3.5, respectively. Coincident with the prolongation of the SH interval at rapid pacing rates, draflazine and ITU + EHNA increased the epicardial fluid adenosine concentrations by 2.2- and 2.6-fold, respectively. In contrast, epicardial transudate levels of adenosine do not change in the presence of PD81,723. The AV nodal effects of draflazine, ITU, EHNA, and PD81,723 were reversed by the A1 adenosine receptor antagonist 8-cyclopentyltheophylline and adenosine deaminase, implicating endogenous adenosine acting at the A1 adenosine receptor. CONCLUSIONS: Adenosine-regulating agents that act in an event- and site-specific manner represent a novel drug design strategy that may potentially be valuable for the long-term treatment of supraventricular arrhythmias and control of ventricular rate during atrial fibrillation or flutter.

Angiotensin II regulation of intracellular calcium in astroglia cultured from rat hypothalamus and brainstem.

This study examines the angiotensin II (Ang II) regulation of intracellular free calcium concentration ([Ca2+]i) in astroglia cultured from the hypothalamus and brainstem of the adult rat. Bath perfusion or rapid puffer application of angiotensin II (Ang II) (1-100 nM) increased [Ca2+]i in both polygonal and stellate astroglia when measured using fura-2 imaging fluorescence microscopy. Ang II increased [Ca2+]i in 96.1 and 95.6% of the polygonal and stellate glial cells, respectively. In normal Tyrode's solution (containing 2 mM CaCl2), the Ang II-stimulated increase in [Ca2+]i characteristically showed a biphasic response, i.e., an initial rapid transient peak followed by a sustained, steady-state plateau of free Ca2+. In both cell types, the selective Ang II type 1 receptor subtype (AT1) antagonist losartan (1 microM) inhibited the Ang II-stimulated increase in [Ca2+]i. The selective AT2 antagonist PD 123319 (1 microM) did not inhibit the Ang II-stimulated increase in [Ca2+]i in either cell type. To define the sources of Ca2+ that participate in the Ang II-stimulated increase in [Ca2+]i in astroglia, experiments were performed in a nominally Ca(2+)-free Tyrode's solution. In either cell type, this resulted in only an initial transient increase of Ca2+ and no sustained plateau of Ca2+ when challenged with Ang II. Thapsigargin (5 microM), cyclopiazonic acid (10 microM), and ryanodine (10 microM), but not caffeine (1-10 mM), inhibited the initial rise in [Ca2+]i. The plateau increase of [Ca2+]i caused by Ang II (100 nM) was reversibly inhibited by both cadmium (100 microM) and nifedipine (10 microM); in contrast, gadolinium (100 microM) had no effect on the plateau increase of [Ca2+]i. These results indicate that Ang II, in physiological concentrations, can activate AT1 receptors to stimulate both Ca2+ release from intracellular stores and Ca2+ influx from the extracellular space to increase [Ca2+]i of polygonal and stellate astroglia.

Effects of intravenous anesthetics on atrial wavelength and atrioventricular nodal conduction in guinea pig heart. Potential antidysrhythmic properties and clinical implications.

BACKGROUND: Supraventricular tachydysrhythmias such as atrial fibrillation frequently complicate the perioperative period. Two electrophysiologic factors critical to the pathogenesis of supraventricular tachydysrhythmias are: 1) atrial wavelength, the product of atrial conduction velocity (CV) and effective refractory period (ERP), and 2) atrioventricular nodal conduction. Modulation of these factors by drugs has important clinical ramifications. The authors studied the effects of propofol, thiopental, and ketamine on atrial wavelength and atrioventricular nodal function in guinea pig isolated atrial trabeculae and hearts, respectively. METHODS: Electrocardiogram recordings in superfused atrial tissue were obtained using hanging microelectrodes. A stimulating and two recording electrodes were placed on a single atrial trabecula, and the interelectrode distance was measured. Atrial ERP determinations were made using a premature stimulus protocol. The time (t) required for a propagated impulse to traverse the interelectrode distance (d) was measured. Conduction velocity was calculated as d/t. Langendorff-perfused guinea pig hearts were instrumented for low atrial pacing (cycle length = 300 ms) and for measurements of stimulusto-His bundle interval, an index of atrioventricular nodal conduction. To investigate the frequency-dependent behavior of the atrioventricular node, computer-based measurements were made of Wenckebach cycle length (WCL) and atrioventricular nodal ERP. RESULTS: Thiopental significantly prolonged atrial ERP in a concentration-dependent manner, whereas propofol and ketamine had no significant effect on atrial refractoriness. In contrast, ketamine caused a dose-dependent decrease in atrial CV, but propofol and thiopental had no significant effect on CV. Therefore, thiopental, ketamine, and propofol caused an increase, a decrease, and no change, respectively, in atrial wavelength. All anesthetics caused a concentration-dependent prolongation of the stimulus-to-His bundle interval, atrioventricular nodal ERP, and WCL. However, on an equimolar basis, significant differences in potencies were found. The concentrations of drug that caused a 20% increase in ERP (ERP20) and WCL (WCL20) for propofol, thiopental, and ketamine were 14 +/- 2 microM, 26 +/- 3 microM, and 62 +/- 11 microM, and 17 +/- 2 microM, 50 +/- 1 microM, and 123 +/- 19 microM (mean +/- SEM), respectively. Therefore, the rank order of potency for frequency-dependent atrioventricular nodal effects is propofol > thiopental > ketamine. CONCLUSION: The authors' results indicate that propofol would be most effective at filtering atrial impulses during supraventricular tachydysrhythmias, whereas thiopental would be most effective at preventing atrial reentrant dysrhythmias. In contrast, ketamine may be most likely to promote atrial reentry while having minimal effect on atrioventricular nodal conduction.

Alterations in rat interlobar artery membrane potential and K+ channels in genetic and nongenetic hypertension.

The renal vasculature plays an important role in the control of blood pressure. K+ channels have been demonstrated to regulate smooth muscle membrane potential and thereby control smooth muscle tone. However, few data are available on K+ channel function in the renal vasculature of hypertensive animals. This study details changes in K+ currents and membrane potential in genetic and nongenetic models of hypertension. The patch-clamp technique and Ca(2+)-imaging fluorescence were used to examine the differences in Wistar-Kyoto (WKY), Sprague-Dawley (SD), spontaneously hypertensive (SHR), and deoxycorticosterone acetate (DOCA) hypertensive single cells of rat kidney interlobar arteries. In current-clamp experiments, SHR and DOCA hypertensive cells were approximately 20 mV more depolarized than the control cells. In voltage-clamp experiments with 4-amino-pyridine and niflumic acid present to inhibit voltage-dependent K+ (K(v)) and Ca(2+)-activated CI- (CI(Ca)) currents, SHR and DOCA hypertensive Ca(2+)-activated K+ (K(Ca)) currents were significantly larger and activated at more negative potentials than the control. Conversely, with charybdotoxin and niflumic acid present to inhibit K(Ca) and CI(Ca) currents, SHR and DOCA hypertensive K(v) current was significantly smaller than the control. Finally, basal and angiotensin II-stimulated peak intracellular free [Ca2+] was greater in the SHR and DOCA hypertensive cells compared with control cells. These results suggest that membrane potential and the activity of K(Ca) and K(v) channels are altered in hypertensive rat renal interlobar arteries and may play a role in the regulation of renal blood flow under physiological and patho-physiological conditions.

Angiotensin II type 2 receptor-mediated regulation of rat neuronal K+ channels.

We have previously shown that angiotensin II (Ang II), via AT2 receptors, increases whole-cell K+ current in cultured rat hypothalamus and brain stern neurons. We have now investigated the AT2 receptor-mediated effects of Ang II on the activity of single delayed rectifier K+ channels in cell-attached membrane patches. In control recordings (bath, 5.4 mmol/L K+; pipette, 140 mmol/L K+), two voltage-dependent channels were recorded with conductances of 34 +/- 4 and 56 +/- 6 pS, respectively (n = 6). When patches were excised, the channels reversed near a membrane potential expected for a K+ channel. In cell-attached patches (-40 mV), Ang II (100 nmol/L) increased open probability of the 56-pS K+ channel from 0.03 +/- 0.01 to 0.21 +/- 0.05 (n = 3). The selective AT2 receptor antagonist PD 123319 (1 mumol/L) but not the AT1 receptor antagonist losartan (1 mumol/L) blocked the actions of Ang II (n = 3). The selective AT2 receptor agonist CGP 42112 (100 nmol/L) produced similar effects to Ang II. Kinetic analysis of the Ang II effect showed that open-time histograms were best fit by two exponential functions. Ang II increased both open-time constants relative to control (control, tau 1 = 0.9 +/- 0.1 milliseconds, tau 2 = 2.3 +/- 0.3 milliseconds; Ang II, tau 1 = 3.1 +/- 0.4 milliseconds, tau 2 = 12.1 +/- 2.4 milliseconds), and PD 123319 blocked this effect (n = 3). The closed-time histogram was not affected by Ang II PD 123319, or losartan. These results suggest that activation of AT2 receptors modulates rat hypothalamus and brain stern neuronal whole-cell K+ current by increasing the open probability of a 56-pS K+ channel.

Voltage-dependent chloride channels: invertebrates to man.

Chloride channels are ubiquitous proteins found in invetebrates to man. Cl- is one of the most abundant biological anions and accounts for a measurable fraction of the electrical conductance of many biological membranes. Physiologically this contributes to cellular processes, including pH regulation, volume regulation, generation of the resting membrane potential, and regulation of membrane excitability. The unitary conductance of voltage-dependent Cl- channels is as diverse as the number of different types of Cl- channels described ranging from 5-450 pS. Cl- channels are highly anion selective passing at least ten anionic species, including all of the halides. Cl- channels are blocked by various agents, including aromatic acids, inorganic cations, and protons. Maintaining high resting conductance and normal excitability, regulating cell volume, and modulating hormone action are some examples of the functions of Cl- channels. Despite the large amount of data accumulated on voltage-dependent Cl- channels, identifying subsets within this class of channels with coherent biophysical features that subserve each specific function is still not possible. At present, the molecular structure for every type of functional Cl- channels has not been determined, but future identification of cloned Cl- channel structures should provide a clearer understanding of the functional properties of background Cl- channels.

Frequency-dependent effects of propofol on atrioventricular nodal conduction in guinea pig isolated heart. Mechanism and potential antidysrhythmic properties.

BACKGROUND: The use of propofol has been associated with episodes of bradycardias. The mechanism(s) underlying these phenomena are not well defined. Therefore we investigated (1) the chronotropic and dromotropic effects of propofol, (2) the frequency-dependent effects of propofol on the atrioventricular (AV) node, and (3) the physiologic mechanism(s) underlying propofol's effects on AV nodal conduction. METHODS: Guinea pig isolated, perfused hearts were instrumented for measurement of atrial rate and AV nodal conduction time in spontaneously beating hearts, or stimulus-to-His bundle (S-H) intervals in atrially paced hearts. In addition, the Wenckebach cycle length, effective refractory period and S-H interval prolongation to an abrupt increase in pacing rate were measured to further define propofol's dromotropic effects and frequency-dependent behavior. RESULTS: Propofol, in a concentration-dependent manner, (1) slowed atrial rate and AV nodal conduction time in spontaneously beating hearts, (2) prolonged the S-H interval in atrially paced hearts, and (3) prolonged Wenckebach cycle length and effective refractory period. The negative dromotropic effect of propofol was greater during atrial pacing than in spontaneously beating hearts. Furthermore, this effect was enhanced at faster pacing rates, indicating frequency-dependent behavior. Atropine significantly antagonized propofol-induced S-H interval prolongation. The results of competition binding studies also supported a M2-muscarinic receptor-mediated mechanism. CONCLUSIONS: We conclude that in the isolated guinea pig heart, propofol slows atrial rate and depresses AV nodal conduction in a concentration-dependent manner. The negative dromotropic effect of propofol shows frequency dependence and is predominantly mediated by M2-muscarinic receptors. Given the marked rate dependence of propofol's AV nodal actions, this anesthetic agent may impart antidysrhythmic protection to those patients susceptible to supraventricular tachycardias.