Molecular pathways underlying the modulation of T-type calcium channels by neurotransmitters and hormones

Low-voltage-activated T-type calcium channels are expressed in various tissues, especially in the brain, where they promote neuronal firing and are involved in slow wave sleep and absence epilepsy. While the transduction pathways by which hormones and neurotransmitters modulate high-voltage-activated calcium channels are beginning to be unraveled, those implicated in T-type calcium channel regulation remain obscure. Several neurotransmitters and hormones regulate native T-type calcium channels, although some contradictory data have been reported depending on the cell type studied. This review focuses on the short-term (minutes range) modulation of T-type calcium channels by neurotransmitters and hormones and on the roles of G proteins and protein kinases in these modulatory effects. Results obtained in different native tissues are discussed and compared with the more recent studies of the three cloned T-type calcium channels CaV3.1, CaV3.2 and CaV3.3 in expression systems.     

Potential role of the glial water channel aquaporin-4 in epilepsy

Recent studies have implicated glial cells in novel physiological roles in the CNS, such as modulation of synaptic transmission, so it is possible that glial cells might have a functional role in the hyperexcitability that is characteristic of epilepsy. Indeed, alterations in distinct astrocyte membrane channels, receptors and transporters have all been associated with the epileptic state. This paper focuses on the potential roles of the glial water channel aquaporin-4 (AQP4) in modulating brain excitability and in epilepsy. We review studies of seizure phenotypes, K(+) homeostasis and extracellular space physiology of mice that lack AQP4 (AQP4(-/-) mice) and discuss the human studies demonstrating alterations of AQP4 in specimens of human epilepsy tissue. We conclude with new studies of AQP4 regulation by seizures and discuss its potential role in the development of epilepsy (epileptogenesis). Although many questions remain unanswered, the available data indicate that AQP4 and its molecular partners might represent important new therapeutic targets.     

Neurotransmitters in alcoholism: A review of neurobiological and genetic studies

Recent advances in the study of alcoholism have thrown light on the involvement of various neurotransmitters in the phenomenon of alcohol addiction. Various neurotransmitters have been implicated in alcohol addiction due to their imbalance in the brain, which could be either due to their excess activity or inhibition. This review paper aims to consolidate and to summarize some of the recent papers which have been published in this regard. The review paper will give an overview of the neurobiology of alcohol addiction, followed by detailed reviews of some of the recent papers published in the context of the genetics of alcohol addiction. Furthermore, the author hopes that the present text will be found useful to novices and experts alike in the field of neurotransmitters in alcoholism.     

Synaptic transmission: exciting times for presynaptic receptors.

Presynaptic receptors play an important role - typically inhibitory--in modulating the strength of synaptic transmission in the brain. Recent studies now reveal that neurotransmitters can enhance synaptic strength by activating ligand-gated ion channels in presynaptic nerve endings.     

KCNQ5, a novel potassium channel broadly expressed in brain, mediates M-type currents

KCNQ2 and KCNQ3, both of which are mutated in a type of human neonatal epilepsy, form heteromeric potassium channels that are expressed in broad regions of the brain. The associated current may be identical to the M-current, an important regulator of neuronal excitability. We now show that the RNA encoding the novel KCNQ5 channel is also expressed in brain and in sympathetic ganglia where it overlaps largely with KCNQ2 and KCNQ3. In addition, it is expressed in skeletal muscle. KCNQ5 yields currents that activate slowly with depolarization and can form heteromeric channels with KCNQ3. Currents expressed from KCNQ5 have voltage dependences and inhibitor sensitivities in common with M-currents. They are also inhibited by M1 muscarinic receptor activation. A KCNQ5 splice variant found in skeletal muscle displays altered gating kinetics. This indicates a molecular diversity of channels yielding M-type currents and suggests a role for KCNQ5 in the regulation of neuronal excitability.     

Drug resistance in epilepsy and the ABCB1 gene: The clinical perspective

Multidrug resistance is one of the most serious problems in the treatment of epilepsy that is likely to have a complex genetic and acquired basis. Various experimental data support the hypothesis that over-expression of antiepileptic drug (AED) transporters may play a pivotal role in drug resistance. Hyyt 6however, key questions concerning their functionality remain unanswered. The idea that P-glycoprotein, encoded by the ABCB1 gene, might mediate at least part of the drug resistance was met with both enthusiasm and skepticism. As in oncology, initial optimism has been clouded subsequently by conflicting results. The first study reporting a positive association between genetic variation in the P-glycoprotein and multidrug-resistant epilepsy was published in 2003. Since then, several other genetic association studies have attempted to verify this result. However, taken overall, the role of P-glycoprotein in drug resistance in epilepsy still remains uncertain. We intend to critically review the inherent problems associated with epilepsy pharmacogenetic studies in general and with ABCB1 polymorphisms studies in particular. The lessons learnt from the ABCB1 studies can help us to guide future association genetics studies to investigate AED resistance, and thereby taking us closer to the cherished dream of personalized AED therapy.     

Conservation genetics in a globally changing environment: present problems,paradoxes and future challenges

Despite recent advances in conservation genetics and related disciplines and the growing impact that conservation genetics is having in conservation biology, our knowledge on several key issues in the field is still insufficient. Here we identify some of these issues together with addressing several paradoxes which have to be solved before conservation genetics can face new challenges that are appearing in the transitory phase from the population genetics into the population genomics era. Most of these issues, paradoxes and challenges, like the central dogma of conservation genetics, the computational, theoretical and laboratory experiment achievements and limitations in the conservation genetics field have been discussed. Further knowledge on the consequences of inbreeding and outbreeding depression in wild populations as well as the capacity of small populations to adapt to local environmental conditions is also urgently needed. The integration of experimental, theoretical and applied conservation genetics will contribute to improve our understanding of methodological and applied aspects of conservation genetics.     

Sodium Channel Toxins and Neurotransmitter Release

Voltage-dependent sodium channels (VDSC) are an important class of ion channels in excitable cells, where they are responsible for the generation and conduction of action potential. In addition, the release of neurotransmitters from nerve terminals is influenced by sodium channel activity. The function of VDSC is subject to modulation by various neurotoxins, such as scorpion toxins, which have long been used as tools in the investigation of neurotransmitter release. This opens an interesting perspective concerning modulation of neurotransmission via pharmacological manipulation of sodium channel properties, which can lead to a better understanding of their physiological and pathological roles. Here we briefly review the studies of neurotoxins acting on sodium channels, focusing primarily on the view of the mechanisms of neurotransmitter release.     

Navigating the pitfalls and promise of landscape genetics

The field of landscape genetics has been evolving rapidly since its emergence in the early 2000s. New applications, techniques and criticisms of techniques appear like clockwork with each new journal issue. The developments are an encouraging, and at times bewildering, sign of progress in an exciting new field of study. However, we suggest that the rapid expansion of landscape genetics has belied important flaws in the development of the field, and we add an air of caution to this breakneck pace of expansion. Specifically, landscape genetic studies often lose sight of the fundamental principles and complex consequences of gene flow, instead favouring simplistic interpretations and broad inferences not necessarily warranted by the data. Here, we describe common pitfalls that characterize such studies, and provide practical guidance to improve landscape genetic investigation, with careful consideration of inferential limits, scale, replication, and the ecological and evolutionary context of spatial genetic patterns. Ultimately, the utility of landscape genetics will depend on translating the relationship between gene flow and landscape features into an understanding of long‐term population outcomes. We hope the perspective presented here will steer landscape genetics down a more scientifically sound and productive path, garnering a field that is as informative in the future as it is popular now.     

GABA is an effective immunomodulatory molecule

In recent years, it has become clear that there is an extensive cross-talk between the nervous and the immune system. Somewhat surprisingly, the immune cells themselves do express components of the neuronal neurotransmitters systems. What role the neurotransmitters, their ion channels, receptors and transporters have in immune function and regulation is an emerging field of study. Several recent studies have shown that the immune system is capable of synthesizing and releasing the classical neurotransmitter GABA (γ-aminobutyric acid). GABA has a number of effects on the immune cells such as activation or suppression of cytokine secretion, modification of cell proliferation and GABA can even affect migration of the cells. The immune cells encounter GABA when released by the immune cells themselves or when the immune cells enter the brain. In addition, GABA can also be found in tissues like the lymph nodes, the islets of Langerhans and GABA is in high enough concentration in blood to activate, e.g., GABA-A channels. GABA appears to have a role in autoimmune diseases like multiple sclerosis, type 1 diabetes, and rheumatoid arthritis and may modulate the immune response to infections. In the near future, it will be important to work out what specific effects GABA has on the function of the different types of immune cells and determine the underlying mechanisms. In this review, we discuss some of the recent findings revealing the role of GABA as an immunomodulator.     

Quantitative genetics,version 3.0: where have we gone since 1987 and where are we headed?

The last 20 years since the previous World Congress have seen tremendous advancements in quantitative genetics, in large part due to the advancements in genomics, computation, and statistics. One central theme of this last 20 years has been the exploitation of the vast harvest of molecular markers—examples include QTL and association mapping, marker-assisted selection and introgression, scans for loci under selection, and methods to infer degree of coancestry, population membership, and past demographic history. One consequence of this harvest is that phenotyping, rather than genotyping, is now the bottleneck in molecular quantitative genetics studies. Equally important have been advances in statistics, many developed to effectively use this treasure trove of markers. Computational improvements in statistics, and in particular Markov Chain Monte Carlo (MCMC) methods, have facilitated many of these methods, as have significantly improved computational abilities for mixed models. Indeed, one could argue that mixed models have had at least as great an impact in quantitative genetics as have molecular markers. A final important theme over the past 20 years has been the fusion of population and quantitative genetics, in particular the importance of coalescence theory with its applications for association mapping, scans for loci under selection, and estimation of the demography history of a population. What are the future directions of the field? While obviously important surprises await us, the general trend seems to be moving into higher and higher dimensional traits and, in general, dimensional considerations. We have methods to deal with infinite-dimensional traits indexed by a single variable (such as a trait varying over time), but the future will require us to treat much more complex objects, such as infinite-dimensional traits indexed over several variables and with graphs and dynamical networks. A second important direction is the interfacing of quantitative genetics with physiological and developmental models as a step towards both the gene–phenotype map as well as predicting the effects of environmental changes. The high-dimensional objects we will need to consider almost certainly have most of their variation residing on a lower (likely much lower) dimensional subspace, and how to treat these constraints will be an important area of future research. Conversely, the univariate traits we currently deal with are themselves projections of more complex structures onto a lower dimensional space, and simply treating these as univariate traits can result in serious errors in understanding their selection and biology. As a field, our future is quite bright. We have new tools and techniques, and (most importantly) new talent with an exciting international group of vibrant young investigators who have received their degrees since the last Congress. One cloud for concern, however, has been the replacement at many universities of plant and animal breeders with plant and animal molecular biologists. Molecular tools are now an integral part of breeding, but breeding is not an integral part of molecular biology.     

Ammonium ion enhances the calcium-dependent gating of a mammalian large conductance, calcium-sensitive K+ channel.

We observed that the current amplitude and activation of expressed, mouse brain large conductance, calcium-sensitive K+ channels (BKCa channels) may be reversibly enhanced following addition of low concentrations of the weakly permeant cation NH4+ to the cytoplasmic face of the channel in excised, inside-out membrane patches from HEK 293 cells. Conductance-voltage relations were left-shifted along the voltage axis by addition of NH4Cl in a concentration-dependent manner, with an EC50 of 18.5 mM. Furthermore, this effect was observed in the presence of cytosolic free calcium (approximately 1 microM), but was absent in a cytosolic bath solution containing nominally zero free calcium (e.g.. 5 mM EGTA only), a condition under which these channels undergo largely voltage-dependent gating. Recordings of single BKCa channel events indicated that NH4+ increased the channel open probability of single channel activity approximately 3-fold, but did not alter the amplitude of single channel currents. These findings suggest that the calcium-sensitive gating of mammalian BKCa channels may be modified by other ions present in cytosolic solution.     

Revealing the molecular determinants of neurotoxin specificity for calcium-activated versus voltage-dependent potassium channels

Potassium channel dysfunction underlies diseases such as epilepsy, hypertension, cardiac arrhythmias, and multiple sclerosis. Neurotoxins that selectively inhibit potassium channels, alpha-KTx, have provided invaluable information for dissecting the contribution of different potassium channels to neurotransmission, vasoconstriction, and lymphocyte proliferation. Thus, alpha-KTx specificity comprises an important first step in potassium channel-directed drug discovery for these diseases. Despite extensive functional and structural studies of alpha-KTx-potassium channel complexes, none have predicted the molecular basis of alpha-KTx specificity. Here we show that by minimizing the differences in binding free energy between selective and nonselective alpha-KTx we are able to identify all of the determinants of alpha-KTx specificity for calcium-activated versus voltage-dependent potassium channels. Because these determinants correspond to unique features of the two types of channels, they provide a way to develop more accurate models of alpha-KTx-potassium channel complexes that can be used to design novel selective alpha-KTx inhibitors.     

An emerging view of presynaptic structure from electron microscopic studies

In response to calcium influx, some of the synaptic vesicles in presynaptic terminals fuse rapidly with the presynaptic membrane, allowing fast synaptic transmission. The regulated recycling of synaptic vesicles at the terminals is required for a sustained release of neurotransmitters. Localization of 'ready to be released' vesicles in close vicinities to voltage-gated calcium channels enables the rapid release of neurotransmitters. Thus, recycling vesicles must translocate from the sites of endocytosis to these release sites. However, the sub-cellular organization that supports this local vesicular traffic remains poorly understood. We will review the results of various electron microscopy studies, which have begun to unveil the structure of presynaptic terminals.     

TREK-1, a K+ channel involved in neuroprotection and general anesthesia

TREK-1 is a two-pore-domain background potassium channel expressed throughout the central nervous system. It is opened by polyunsaturated fatty acids and lysophospholipids. It is inhibited by neurotransmitters that produce an increase in intracellular cAMP and by those that activate the Gq protein pathway. TREK-1 is also activated by volatile anesthetics and has been suggested to be an important target in the action of these drugs. Using mice with a disrupted TREK-1 gene, we now show that TREK-1 has an important role in neuroprotection against epilepsy and brain and spinal chord ischemia. Trek1-/- mice display an increased sensitivity to ischemia and epilepsy. Neuroprotection by polyunsaturated fatty acids, which is impressive in Trek1+/+ mice, disappears in Trek1-/- mice indicating a central role of TREK-1 in this process. Trek1-/- mice are also resistant to anesthesia by volatile anesthetics. TREK-1 emerges as a potential innovative target for developing new therapeutic agents for neurology and anesthesiology.     

Molecular population genetics and the search for adaptive evolution in plants

The first papers on plant molecular population genetics were published approximately 10 years ago. Since that time, well over 50 additional studies of plant nucleotide polymorphism have been published, and many of these studies focused on detecting the signature of balancing or positive selection at a locus. In this review, we discuss some of the theoretical and statistical issues surrounding the detection of selection, with focus on plant populations, and we also summarize the empirical plant molecular population genetics literature. At face value, the literature suggests that a history of balancing or positive selection in plant genes is rampant. In two well-studied taxa (maize and Arabidopsis) over 20% of studied genes have been interpreted as containing the signature of selection. We argue that this is probably an overstatement of the prevalence of natural selection in plant genomes, for two reasons. First, demographic effects are difficult to incorporate and have generally not been well integrated into the plant population genetics literature. Second, the genes studied to date are not a random sample, so selected genes may be overrepresented. The next generation of studies in plant molecular population genetics requires additional sampling of local populations, explicit comparisons among loci, and improved theoretical methods to control for demography. Eventually, candidate loci should be confirmed by explicit consideration of phenotypic effects.     

Forty Years of Sodium Channels: Structure,Function, Pharmacology,and Epilepsy

Voltage-gated sodium channels initiate action potentials in brain neurons. In the 1970s, much was known about the function of sodium channels from measurements of ionic currents using the voltage clamp method, but there was no information about the sodium channel molecules themselves. As a postdoctoral fellow and staff scientist at the National Institutes of Health, I developed neurotoxins as molecular probes of sodium channels in cultured neuroblastoma cells. During those years, Bruce Ransom and I crossed paths as members of the laboratories of Marshall Nirenberg and Philip Nelson and shared insights about sodium channels in neuroblastoma cells from my work and electrical excitability and synaptic transmission in cultured spinal cord neurons from Bruce’s pioneering electrophysiological studies. When I established my laboratory at the University of Washington in 1977, my colleagues and I used those neurotoxins to identify the protein subunits of sodium channels, purify them, and reconstitute their ion conductance activity in pure form. Subsequent studies identified the molecular basis for the main functions of sodium channels—voltage-dependent activation, rapid and selective ion conductance, and fast inactivation. Bruce Ransom and I re-connected in the 1990s, as ski buddies at the Winter Conference on Brain Research and as faculty colleagues at the University of Washington when Bruce became our founding Chair of Neurology and provided visionary leadership of that department. In the past decade my work on sodium channels has evolved into structural biology. Molecular modeling and X-ray crystallographic studies have given new views of sodium channel function at atomic resolution. Sodium channels are also the molecular targets for genetic diseases, including Dravet Syndrome, an intractable pediatric epilepsy disorder with major co-morbidities of cognitive deficit, autistic-like behaviors, and premature death that is caused by loss-of-function mutations in the brain sodium channel Na_V1.1. Our work on a mouse genetic model of this disease has shown that its multi-faceted pathophysiology and co-morbidities derive from selective loss of electrical excitability and action potential firing in GABAergic inhibitory neurons, which disinhibits neural circuits throughout the brain and leads directly to the epilepsy, premature death and complex co-morbidities of this disease. It has been rewarding for me to use our developing knowledge of sodium channels to help understand the pathophysiology and to suggest potential therapeutic approaches for this devastating childhood disease.     

Ion channels and postsynaptic potentials

Neurotransmitters open transmembrane ion channels in target membranes. At the motor endplate, the open time of channels activated by acetylcholine determines the time course of the endplate current. Endplate channels are selective for cations and their conductance and open times are influenced by the nature of the permeating cation. Similar effects have been seen at anion-selective inhibitory synapses. Some 'blocking' drugs that depress postsynaptic responses to neurotransmitters produce effects not consistent with a simple model in which they block open channels. Channels activated by neurotransmitters can show subconductance states that may reflect the fundamental mode of operation of a variety of ion channels.     

Neurotensin analogs indications for use as potential antipsychotic compounds

This review will be an update, focusing on the central nervous system (CNS) roles of the neurotransmitter, neurotensin. We will provide a summary of current knowledge about neurotensin, why it is an important peptide to study, and where the field is heading. Special emphasis is placed on the development of neurotensin analogs, which has been a major effort of our group, the potential role of neurotensin in Parkinson's disease, and the interaction of neurotensin with other neurotransmitters as evidenced by microdialysis studies.