Potassium Channels

Perforated whole-cell patch-clamp recordings obtained with nystatin are frequently used to preserve intracellular integrity. However, the perforated-patch configuration may sometimes undergo a spontaneous change into the conventional whole-cell configuration, especially when lymphocytes are investigated. The electrophysiological criteria-- previously described--for establishing the existence of the perforated whole-cell configuration have been shown to be insufficient. Thus, the dye eosin, applied to the pipette solution, was tested as a tool for discriminating between the perforated and the conventional whole-cell configurations on rat T-lymphocytes. The dye never entered the cell from the pipette during the entire measurement in the perforated whole-cell configuration. In contrast, all cells in the conventional whole-cell configuration became red immediately after membrane rupture. Eosin barely changed the currents studied. The results suggest that eosin is a dye of choice for verifying a true perforated-patch configuration.

Metabotropic glutamate receptors (mGluRs) can increase intracellular Ca2+ concentration via Ins(1,4,5)P3- and ryanodine-sensitive Ca2+ stores in neurons. Both types of store are coupled functionally to Ca2+-permeable channels found in the plasma membrane. The mGluR-mediated increase in intracellular Ca2+ concentration can activate Ca2+-sensitive K+ channels and Ca2+-dependent nonselective cationic channels. These mGluR-mediated effects often result from mobilization of Ca2+ from ryanodine-sensitive, rather than Ins(1,4,5)P3-sensitive, Ca2+ stores, suggesting that close functional interactions exist between mGluRs, intracellular Ca2+ stores and Ca2+-sensitive ion channels in the membrane.

Hyperkalemic periodic paralysis (HyperKPP) is a disease characterized by periods of myotonic discharges and paralytic attacks causing weakness, the latter associated with increases in plasma [K(+)]. The myotonic discharge is due to increased Na(+) influx through defective Na(+) channels that triggers generation of several action potentials. The subsequent increase in extracellular K(+) concentration causes excessive membrane depolarization that inactivates Na(+) channels triggering the paralysis. None of the available treatments is fully effective. This paper reviews the capacity of Na(+) K(+)ATPase pumps, KATP and ClC-1 Cl(-) channels in improving membrane excitability during muscle activity and how using these three membrane components we can study future and more effective treatments for HyperKPP patients. The review of current patents related to HyperKPP reinforces the need of novel approaches for the treatment of this channelopathy.

Abstract: The peptide neurotransmitter Phe-Met-Arg-PheNH2 (FMRFamide) increases outward K+ currents and promotes dephosphorylation of many phosphoproteins in Aplysia sensory neurons. We examined FMRFamide-induced current responses in sensory neurons injected with thiophosphorylated protein phosphate inhibitor-1 and inhibitor-2 (I-1 and I-2), two structurally different vertebrate protein phosphatase-1 (PP1) inhibitors to define a role for PP1 in the physiological actions of FMRFamide. Thiophosphorylated I-1 and I-2 both reduced the amplitude of outward currents elicited by FMRFamide by 50–60% and were as effective as microcystin-LR, which inhibited both PP1 and protein phosphatase-2A in Aplysia neuronal extracts. These data suggested that of the two major neuronal protein serine/threonine phosphatases, FMRFamide utilized primarily PP1 to open serotonin-sensitive K+ (S-K+) channels. Earlier studies showed that a membrane-associated phosphatase regulated S-K+ channels in cell-free patches from sensory neurons. Utilizing its unique substrate specificity and inhibitor sensitivity, we have characterized PP1 as the principal protein phosphatase associated with neuronal plasma membranes. Two protein phosphatase activities (apparent Mr values of 170,000 and 38,000) extracted from crude membrane preparations from the Aplysia nervous system were shown to be isoforms of PP1. These biochemical and physiological studies suggest that PP1 is preferentially associated with neuronal membranes and that its activity may be required for the induction of outward K+ currents in the Aplysia sensory neurons by FMRFamide.

Synapses are specialised structures where interneuronal communication takes place. Not only brain function is absolutely dependent on synaptic activity, but also most of our organs are intimately controlled by synaptic activity. Synapses re therefore an ideal target to act upon and poisonous species have evolved fascinating neurotoxins capable of shutting down neuronal communication by blocking or activating essential components of the synapse. By hijacking key proteins of the communication machinery, neurotoxins are therefore extremely valuable tools that have, in turn, greatly helped our understanding of synaptic biology. Moreover, analysis and understanding of the molecular strategy used by certain neurotoxins has allowed the design of entirely new classes of drugs acting on specific targets with high selectivity and efficacy. This chapter will discuss the different classes of marine neurotoxins, their effects on neurotransmitter release and how they act to incapacitate key steps...

Potassium (K+) channels are the most heterogeneous and widely distributed class of ion channels. K(+) channels are dynamic pore-forming transmembrane proteins known to play important roles in all cell types underlying both normal and pathophysiological functions. Essential for such diverse physiological processes as nerve impulse propagation, muscle contraction, cellular activation and the secretion of biologically active molecules, various K(+) channels are recognized as potential therapeutic targets in the treatment of multiple sclerosis, Alzheimer's disease, Parkinson's disease, epilepsy, stroke, brain tumors, brain/spinal cord ischemia, pain and schizophrenia, migraine, as well as cardiac arrhythmias, pulmonary hypertension, diabetes, cervical cancer, urological diseases and sepsis. In addition to their importance as therapeutic targets, certain K(+) channels are gaining attention for their beneficial roles in anesthesia, neuroprotection and cardioprotection. The K(+) ch...

Long QT syndrome is a disease of delayed ventricular repolarization. It manifests clinically as recurrent syncope and sudden cardiac death caused by an atypical form of polymorphic ventricular tachycardia known as torsades de pointes (TdP). Evidence obtained from the studies using the rabbit left and right ventricular wedge preparations indicates that the development of TdP is relying not only on the genesis of an R-on-T trigger, but also on the formation of a functional reentrant substrate. When ventricular endocardial or subendocardial repolarization is prolonged either because of gene mutations or by drugs that reduce the net repolarization current, cell membrane potential fluctuates during phase 2 of the action potential phase 2 because of reactivation of L-type calcium current, that is, the appearance of phase 2 early afterdepolarization (EAD). In the rabbit left ventricular wedge, QT prolongation and EAD due to pure IKr inhibition are accompanied by a disproportional increase in transmural dispersion of repolarization (TDR). Early afterdepolarization in endocardium or subendocardium is able to produce new action potentials in cells with a relatively short action potential duration (eg, ventricular epicardium) probably via an electrotonic effect when TDR is large enough. This, in turn, results in an R-on-T extrasystole that is capable of initiating TdP. Enhanced TDR is essential not only for the genesis of the first initiating beat of TdP by facilitating the propagation of EAD, but also for the maintenance of TdP by serving as a functional reentrant substrate.

Trimebutine [2-dimethylamino-2-phenylbutyl-3,4,5-trimethoxybenzoate hydrogen maleate (TMB)] has been demonstrated to be active for relieving abdominal pain in humans. To better understand its mechanism of action, we have tested TMB; nor-TMB, its main metabolite in humans; and their respective stereoisomers for their affinity toward sodium channels labeled by [3H]batrachotoxin, their effect on sodium, potassium, and calcium currents in rat dorsal root ganglia neurons, and their effect on veratridine-induced glutamate release from rat spinal cord slices. TMB has also been tested in an animal model of local anesthesia. TMB (Ki = 2.66 +/- 0.15 microM) and nor-TMB (Ki = 0.73 +/- 0.02 microM) displaced [3H]batrachotoxin from its binding site with affinities similar to that of bupivacaine (Ki = 7.1 +/- 0.9 microM). nor-TMB was found to block veratridine-induced glutamate release with an IC50 value of 8.5 microM, which is very similar to that of bupivacaine (IC50 = 8.2 microM); the effect o...

Diabetes developing within the first 6 months of life is rarely, if ever, caused by a classic type 1 diabetes-related autoimmune process. Currently, patients developing diabetes before 6 months of age are defined as having neonatal diabetes although this terminology possibly needs amending. Neonatal diabetes has a transient and permanent form and over 10 distinct genetic anomalies or mutations have been identified causing the disease. Transient neonatal diabetes can be caused by defects in the normal methylation pattern of an imprinted gene on chromosome 6 and by mutations in the 2 genes encoding the beta-cell ATP-sensitive potassium channel which is vital to normal glucose-stimulated insulin secretion. A genetic cause can be identified in over 90% of transient cases. Permanent neonatal diabetes can be caused by mutations in beta-cell transcription factors leading to abnormal pancreatic development often with other significant developmental anomalies, by defects in the glucose sensi...

Many drugs fail in the clinical trials and therefore do not reach the market due to adverse effects on cardiac electrical function. This represents a growing concern for both regulatory and pharmaceutical agencies as it translates into important socio-economic costs. Drugs affecting cardiac activity come from diverse pharmacological groups and their interaction with cardiac electrophysiology can result in increased risk of potentially life threatening arrhythmias, such as Torsade de Pointes. The mechanisms of drug interaction with the heart are very complex and the effects span from the ion channel to the whole organ level. This makes their investigation using solely experimental in vitro and in vivo techniques very difficult. Computational modelling of cardiac electrophysiological behaviour has provided insight into the mechanisms of cardiac arrhythmogenesis, with high spatio-temporal resolution, from the ion channel to the whole organ level. It therefore represents a powerful tool in investigating mechanisms of drug-induced changes in cardiac behaviour and in their pro-arrhythmic potential. This article presents a comprehensive review of the recent advances in detailed models of drug action on cardiac electrophysiological activity.