Diverse levels of an inwardly rectifying potassium conductance generate heterogeneous neuronal behavior in a population of dorsal cochlear nucleus pyramidal neurons

Leao RM, Li S, Doiron B, Tzounopoulos T. Diverse levels of an inwardly rectifying potassium conductance generate heterogeneous neuronal behavior in a population of dorsal cochlear nucleus pyramidal neurons. J Neurophysiol 107: 3008-3019, 2012. First published February 29, 2012; doi:10.1152/jn.00660.2011.-Homeostatic mechanisms maintain homogeneous neuronal behavior among neurons that exhibit substantial variability in the expression levels of their ionic conductances. In contrast, the mechanisms, which generate heterogeneous neuronal behavior across a neuronal population, remain poorly understood. We addressed this problem in the dorsal cochlear nucleus, where principal neurons exist in two qualitatively distinct states: spontaneously active or not spontaneously active. Our studies reveal that distinct activity states are generated by the differential levels of a Ba2+-sensitive, inwardly rectifying potassium conductance (K-ir). Variability in K-ir maximal conductance causes variations in the resting membrane potential (RMP). Low K-ir conductance depolarizes RMP to voltages above the threshold for activating subthreshold-persistent sodium channels (Na-p). Once Na-p channels are activated, the RMP becomes unstable, and spontaneous firing is triggered. Our results provide a biophysical mechanism for generating neural heterogeneity, which may play a role in the encoding of sensory information.

Phoneutria nigriventer spider toxin Tx2-6 causes priapism and death: A histopathological investigation in mice

Phoneutria nigriventer spider bite causes priapism, an effect attributed to the peptide toxins Tx2-5 and Tx2-6 and involving nitric oxide. Tx2-6 (MW = 5287) is known to delay the inactivation of Sodium channels in the same fashion as many other venom toxins. In the present study we evaluated the i.p. dose that induces priapism and the other symptoms in mice. Animals killed by the toxin or crude venom (0.85 mg/kg) were autopsied and a pathological study of brain, lung, kidney, liver and heart was undertaken using standard techniques. The same protocol was employed with animals injected with crude venom. Results showed that priapism is the first sign of intoxication, followed by piloerection, abundant salivation and tremors. An i.p. injection of about 0.3 mu g/kg induced only priapism with minimal side-effects. The most remarkable histological finding was a general vascular congestion in all organs studied. Penis showed no necrosis or damage. Lungs showed vascular congestion and alveolar hemorrhage. Heart showed also sub-endothelial hemorrhage. Brain showed only a mild edema and vascular congestion. Results obtained with crude venom closely resemble those of purified toxin. We conclude that Tx2-6 have profound effects on the vascular bed especially in lungs and heart, which may be the cause of death. Interestingly brain tissue was less affected and the observed edema may be attributed to respiratory impairment. To the best of our knowledge this is the first histopathological investigation on this toxin and venom suggesting a possible cause of death. (C) 2012 Elsevier Ltd. All rights reserved.

The occurrence of defensive alkaloids in non-integumentary tissues of the Brazilian red-belly toad Melanophryniscus simplex (Bufonidae)

The red-belly toads (Melanophryniscus) of southern South America secrete defensive alkaloids from dermal granular glands. To date, all information on Melanophryniscus alkaloids has been obtained by extraction from either skins or whole organisms; however, in other amphibians, tetrodotoxins, samandarines, and bufadienolides have been detected in both skin and other organs, which raise the possibility that lipophilic alkaloids may occur in non-integumentary tissues in Melanophryniscus as well. To test this hypothesis, we studied the distribution of alkaloids in the skin, skeletal muscle, liver, and mature oocytes of the red-belly toad M. simplex from three localities in southern Brazil. Gas chromatography and mass spectrometry of skin extracts from 11 individuals of M. simplex resulted in the detection of 47 alkaloids (including isomers), 9 unclassified and 38 from 12 known structural classes. Each alkaloid that was present in the skin of an individual was also present in the same relative proportion in that individual's skeletal muscle, liver, and oocytes. The most abundant and widely distributed alkaloids were the pumiliotoxins 251D, 267C, and 323A, 5,8-disubstituted indolizidines 207A and 223D, 5,6,8-trisubstituted indolizidine 231B, 3,5-disubstituted pyrrolizidines cis-223B and cis- and trans-251K, and izidine 211C. We report the first record of piperidines in Melanophryniscus, bringing the total number of alkaloid classes detected in this genus to 16. Alkaloid composition differed significantly among the three study sites. The functional significance of defensive chemicals in non-integumentary tissues is unknown.

Signalling mechanisms affecting regulated neurotrophin secretion in hippocampal neurons

Die Neurotrophine aus Säugetiere BDNF und NT-3 sind von Neuronen sekretierte Wachstumsfaktoren. Ferner sind Neurotrophine in verschiedene Formen der aktivitätsabhängigen synaptische Plastizität involviert. Obwohl die Ausschüttung von Neurotrophine aus Synapsen beschrieben worden ist, sind die intrazellulären Signalkaskaden, die die synaptische Ausschüttung von Neurotrophine regulieren, bei weitem nicht verstanden. Deswegen ist die Analyse der Sekretion von Neurotrophine auf subzellulärer Ebene erforderlich, um die genaue Rolle von präsynaptische und postsynaptische NT-Sekretion in der synaptischen Plastizität aufzudecken. In der vorliegenden Arbeit wurden die Kulturen von dissoziierten hippocampalen Neuronen aus Ratten mit grün fluoreszierenden Protein-markierten Konstrukten von BDNF und NT-3 transfiziert und Neurotrophine-enthaltenden Vesikeln durch die Colokalisierung mit dem cotransfizierten postsynaptischen Marker PSD-95-DsRed an glutamatergen Synapsen identifiziert. Depolarisationsinduzierte Sekretion von BDNF und NT-3 wurde per Direktaufnahme am Fluoreszenzmikroskop beobachtet. Die unvermittelte postsynaptische Depolarisation mit erhöhtem Kalium, in Gegenwart von Inhibitoren der synaptischen Transmission, erlaubte die Untersuchung der Signalwege, die am postsynaptischen Sekretionsprozess der Neurotrophinvesikel beteiligt sind. Es konnte gezeigt werden, dass die depolarisationsinduzierte postsynaptische Ausschüttung der Neurotrophine durch Calcium-Einstrom ausgelöst wird, entweder über L-Typ-spannungsabhängige Calcium-Kanäle oder über NMDA-Rezeptoren. Eine anschließende Freisetzung von Calcium aus intrazellulären Speichern über Ryanodin-Rezeptoren ist für den Sekretionsprozess erforderlich. Die postsynaptische Neurotrophinausschüttung wird durch KN-62 und KN-93 gehemmt, was auf eine unmittelbare Abhängigkeit von aktiver alpha-Calcium-Calmodulin-abhängige Proteinkinase II (CaMKII) hinweist. Der Inhibitor der cAMP/Proteinkinase A (PKA), Rp-cAMP-S, sowie der NO-Donor, SNP, minderten die Neurotrophinausschüttung. Hingegen blieben die Erhöhung des intrazellulären cAMP und der NO-Synthase-Inhibitor L-NMMA ohne Wirkung. Mit dem Trk-Inhibitor K252a konnte gezeigt werden, dass autokrine Neurotrophin-induzierte Neurotrophinausschüttung nicht an der synaptischen Freisetzung der Neurotrophine beiträgt und, dass BDNF seine eigene postsynaptische Sekretion nicht auslöst. Freisetzungsexperimente mit dem Fluoreszenz-Quencher Bromphenolblau konnten den Nachweis erbringen, dass asynchrone und anhaltende Fusionsporenöffnung von Neurotrophinvesikeln während der Sekretion stattfindet. Wegen der im Vergleich zum komplexen Sekretionsprozess schnellen Fusionsporenöffnung, scheint die Freisetzungsgeschwindigkeit von Neurotrophine durch ihre Diffusion aus dem Vesikel begrenzt. Zusammenfassend zeigen diese Ergebnisse eine starke Abhängigkeit der aktivitätsabhängigen postsynaptischen Neurotrophinausschüttung vom Calcium-Einstrom, von der Freisetzung von Calcium aus internen Speichern, von der Aktivierung der CaMKII und einem intakten Funktion der PKA, während der Trk-Signalweg, die Aktivierung von Natrium-Kanäle und NO-Signale nicht erforderlich sind.

Neuronal expression of the ubiquitin ligase Nedd4-2 in rat dorsal root ganglia: modulation in the spared nerve injury model of neuropathic pain

Neuronal hyperexcitability following peripheral nerve lesions may stem from altered activity of voltage-gated sodium channels (VGSCs), which gives rise to allodynia or hyperalgesia. In vitro, the ubiquitin ligase Nedd4-2 is a negative regulator of VGSC α-subunits (Na(v)), in particular Na(v)1.7, a key actor in nociceptor excitability. We therefore studied Nedd4-2 in rat nociceptors, its co-expression with Na(v)1.7 and Na(v)1.8, and its regulation in pathology. Adult rats were submitted to the spared nerve injury (SNI) model of neuropathic pain or injected with complete Freund's adjuvant (CFA), a model of inflammatory pain. L4 dorsal root ganglia (DRG) were analyzed in sham-operated animals, seven days after SNI and 48 h after CFA with immunofluorescence and Western blot. We observed Nedd4-2 expression in almost 50% of DRG neurons, mostly small and medium-sized. A preponderant localization is found in the non-peptidergic sub-population. Additionally, 55.7 ± 2.7% and 55.0 ± 3.6% of Nedd4-2-positive cells are co-labeled with Na(v)1.7 and Na(v)1.8 respectively. SNI significantly decreases the proportion of Nedd4-2-positive neurons from 45.9 ± 1.9% to 33.5 ± 0.7% (p<0.01) and the total Nedd4-2 protein to 44% ± 0.13% of its basal level (p<0.01, n=4 animals in each group, mean ± SEM). In contrast, no change in Nedd4-2 was found after peripheral inflammation induced by CFA. These results indicate that Nedd4-2 is present in nociceptive neurons, is downregulated after peripheral nerve injury, and might therefore contribute to the dysregulation of Na(v)s involved in the hyperexcitability associated with peripheral nerve injuries.

NaV1.5 and interacting proteins in human arrhythmogenic cardiomyopathy

Evaluation of: Noorman M, Hakim S, Kessler E et al. Remodeling of the cardiac sodium channel, connexin43, and plakoglobin at the intercalated disk in patients with arrhythmogenic cardiomyopathy. Heart Rhythm 10(3), 412-419 (2013). Arrhythmogenic cardiomyopathy (AC) is a heart muscle disease characterized by a progressive replacement of the ventricular myocardium with adipose and fibrous tissue. This disease is often associated with mutations in genes encoding desmosomal proteins in the majority of patients. Based on results obtained from recent experimental models, a disturbed distribution of gap junction proteins and cardiac sodium channels may also be observed in AC phenotypes, secondary to desmosomal dysfunction. The study from Noorman et al. examined heart sections from patients diagnosed with AC and performed immunohistochemical analyses of N-cadherin, PKP2, PKG, Cx43 and the cardiac sodium channel NaV1.5. Altered expression/distribution of Cx43, PKG and NaV1.5 was found in most cases of patients with AC. The altered expression and/or distribution of NaV1.5 channels in AC hearts may play a mechanistic role in the arrhythmias leading to sudden cardiac death in AC patients. Thus, NaV1.5 should be considered as a supplemental element in the evaluation of risk stratification and management strategies. However, additional experiments are required to clearly understand the mechanisms leading to AC phenotypes.

Rhythmic synaptic activity induced by mechanical injury of rat CA3 hippocampal area.

Mechanical injury of the CNS frequently results from accidents but also occurs in the course of neurosurgical interventions. A great variety of anatomical and physiological changes have been described to evolve after a brain trauma yet only little is known about processes that occur during a trauma. In the present study, I obtained whole-cell patch clamp recordings from pyramidal cells in hippocampal slice cultures while mechanically lesioning the CA3 area. Electrophysiological analysis revealed that traumatic injury massively increased excitatory and inhibitory synaptic activity in the entire CA3 region. Cutting the CA3 region induced highly rhythmic excitatory postsynaptic currents (EPSCs) that reached frequencies of around 70 Hz. Blocking voltage-dependent sodium channels with tetrodotoxin prevented the increase in synaptic activity and injury-induced neurotransmitter release in CA3 remote from the lesion site. With fast synaptic transmission blocked only neurons in the immediate vicinity of a lesion depolarized and fired action potentials upon mechanical damage. I hence suggest that mechanical injury damages the membrane and induces action potential firing in only a small population of neurons. This activity is then propagated throughout the undamaged CA3 network inducing highly rhythmic discharges. Thus mechanical brain injury initiates immediate functional changes that exceed the lesion site.

Transporte electrogénico en el colon de ratas privadas de sodio : bloqueo de canales epiteliales versus inhibición de la NA, K-ATPASA

La privación dietaria de sodio estimula la secreción de aldosterona. En el colon de rata, la aldosterona elevada aumenta la absorción de Na+, pero además torna electrogénico el mecanismo de absorción (normalmente electroneutro). Dicho transporte electrogénico puede suprimirse mediante el bloqueo de los canales epiteliales de Na+ en la membrana apical o la inhibición de la Na, KATPasa de la membrana basolateral. La absorción electrogénica de sodio está estrechamente acoplada al metabolismo aerobio, pero se desconoce si el bloqueo de los canales de Na+ reduce el consumo de oxígeno en igual medida que la inhibición de la Na, K-ATPasa. Se obtuvieron preparados de mucosa aislada del colon distal de ratas alimentadas con una dieta hiposódica por 10 días. Se determinó simultáneamente la corriente de cortocircuito y el consumo de oxígeno en condición basal y luego del bloqueo de canales de Na+ con amilorida (n=12) o de la Na, K-ATPasa con uabaína (n=12). Ambos tratamientos redujeron la corriente de cortocircuito en igual medida (>80%), pero la reducción en el consumo de oxígeno fue mayor con uabaína que con amilorida (p<0.03). Esto se debe probablemente a que la Na, KATPasa cumple otras funciones, además del transporte transepitelial de Na+, que son suprimidas por la uabaína pero no por la amilorida.

Cocaine reward models: Conditioned place preference can be established in dopamine- and in serotonin-transporter knockout mice

Cocaine and methylphenidate block uptake by neuronal plasma membrane transporters for dopamine, serotonin, and norepinephrine. Cocaine also blocks voltage-gated sodium channels, a property not shared by methylphenidate. Several lines of evidence have suggested that cocaine blockade of the dopamine transporter (DAT), perhaps with additional contributions from serotonin transporter (5-HTT) recognition, was key to its rewarding actions. We now report that knockout mice without DAT and mice without 5-HTT establish cocaine-conditioned place preferences. Each strain displays cocaine-conditioned place preference in this major mouse model for assessing drug reward, while methylphenidate-conditioned place preference is also maintained in DAT knockout mice. These results have substantial implications for understanding cocaine actions and for strategies to produce anticocaine medications.

Identification of genes differentially expressed by nerve growth factor- and neurotrophin-3-dependent sensory neurons

The neurotrophins nerve growth factor (NGF) and neurotrophin-3 (NT3) support the survival of subpopulations of primary sensory neurons with defined and distinct physiological characteristics. Only a few genes have been identified as being differentially expressed in these subpopulations, and not much is known about the nature of the molecules involved in the processing of sensory information in NGF-dependent nociceptive neurons or NT3-dependent proprioceptive neurons. We devised a simple dorsal root ganglion (DRG) explant culture system, allowing the selection of neuronal populations preferentially responsive to NGF or NT3. The reliability of this assay was first monitored by the differential expression of the NGF and NT3 receptors trkA and trkC, as well as that of neuropeptides and calcium-binding proteins. We then identified four differentially expressed sodium channels, two enriched in the NGF population and two others in the NT3 population. Finally, using an optimized RNA fingerprinting protocol, we identified 20 additional genes, all differentially expressed in DRG explants cultured with NGF or NT3. This approach thus allows the identification of large number of genes expressed in subpopulations of primary sensory neurons and opens the possibility of studying the molecular mechanisms of nociception and proprioception.

Expressão dos genes codificadores de canais de sódio Nav 1.7, Nav 1.8 e Nav 1.9 em portadores da Síndrome de Ardência Bucal

A síndrome de ardência bucal (SAB) é uma condição caracterizada pelo sintoma de ardência na mucosa oral, na ausência de qualquer sinal clínico. Sua etiologia é desconhecida e, até o momento, não dispõe de tratamento efetivo. Há entretanto características de doença neuropática que justificam investigações nesse sentido. O objetivo desse estudo foi mensurar a expressão gênica dos receptores de canais de sódio, Nav 1.7, Nav 1.8 e Nav 1.9, nos pacientes portadores de SAB. A casuística foi composta por dois grupos sendo o grupo de estudo composto por 12 pacientes portadores de SAB, selecionados através do critério estabelecido pela International Headache Society, em 2013 e o grupo controle composto por 4 pacientes não portadores de SAB. As amostras analisadas foram coletadas do dorso lingual, por meio de biópsia realizada com punch de 3 mm e profundidade de 3 mm, estas foram submetidas ao método de análise RT-PCR em tempo real. A expressividade dos genes de canais de sódio foi avaliada nos indivíduos portadores de SAB em relação aos do grupo controle, sendo esta calculada a partir da normalização dos dados da quantificação destes com os da expressão do gene constitutivo (GAPDH), pelo método de Cicle Threshold comparativo e analisados estatisticamente por meio do teste estatístico Mann-Whitnney. Observou-se o aumento da expressão gênica do Nav 1.7 (fold-change = 38.70) e diminuição da expressão gênica do Nav 1.9 (fold-change = 0.89), porém sem diferenças estatisticamente significativas entre os grupos analisados. O gene Nav 1.8 não foi expresso em nenhuma das amostras analisadas. O Nav 1.7 expressa-se tanto em neurônios nociceptivos quanto no sistema nervoso autônomo e mutações no Nav 1.9 tem sido associada a perda de percepção dolorosa. Os resultados obtidos embora não estatisticamente significativos são compatíveis com as características da doença, justificando a extensão dos estudos na linha expressão de genes codificadores dos canais de sódio em pacientes com SAB.

Le système endocannabinoïde dans la rétine du singe : expression, localisation et fonctions

Le cannabis produit de nombreux effets psychologiques et physiologiques sur le corps humain. Les molécules contenues dans cette plante, désignées comme « phytocannabinoïdes », activent un système endogène qu’on appelle le système endocannabinoïde (eCB). Les effets de la consommation de cannabis sur la vision ont déjà été décrits sans cependant de formulation sur les mécanismes sous-jacents. Ces résultats comportementaux suggèrent, malgré tout, la présence de ce système eCB dans le système visuel, et particulièrement dans la rétine. Cette thèse vise donc à caractériser l’expression, la localisation et le rôle du système eCB dans la rétine du singe vervet, une espèce animale ayant un système visuel semblable à celui de l’humain. Nous avons mis au point un protocole expérimental d’immunohistochimie décrit dans l’article apparaissant dans l’Annexe I que nous avons utilisé pour répondre à notre objectif principal. Dans une première série de quatre articles, nous avons ainsi caractérisé l’expression et la localisation de deux récepteurs eCBs reconnus, les récepteurs cannabinoïdes de type 1 (CB1R) et de type 2 (CB2R), et d’un 3e présumé récepteur aux cannabinoïdes, le récepteur GPR55. Dans l’article 1, nous avons démontré que CB1R et une enzyme clé de ce système, la fatty acid amide hydrolase (FAAH), sont exprimés dans les parties centrale et périphérique de la rétine, et abondamment présents dans la fovéa, une région où l’acuité visuelle est maximale. Dans l’article 2, nous avons localisé le CB2R dans des cellules gliales de la rétine : les cellules de Müller et nous avons proposé un modèle sur l’action de cette protéine dans la fonction rétinienne faisant appel à une cascade chimique impliquant les canaux potassiques. Dans l’article 3, nous avons observé le GPR55 exclusivement dans les bâtonnets qui sont responsables de la vision scotopique et nous avons soumis un deuxième modèle de fonctionnement de ce récepteur par le biais d'une modulation des canaux calciques et sodiques des bâtonnets. Vu que ces 3 récepteurs se retrouvent dans des cellules distinctes, nous avons suggéré leur rôle primordial dans l’analyse de l’information visuelle au niveau rétinien. Dans l’article 4, nous avons effectué une analyse comparative de l’expression du système eCB dans la rétine de souris, de toupayes (petits mammifères insectivores qui sont sont considérés comme l’étape intermédiaire entre les rongeurs et les primates) et de deux espèces de singe (le vervet et le rhésus). Ces résultats nous ont menés à présenter une hypothèse évolutionniste quant à l’apparition et à la fonction précise de ces récepteurs. Dans les articles subséquents, nous avons confirmé notre hypothèse sur le rôle spécifique de ces trois récepteurs par l’utilisation de l’électrorétinographie (ERG) après injection intravitréenne d’agonistes et d’antagonistes de ces récepteurs. Nous avons conclu sur leur influence indéniable dans le processus visuel rétinien chez le primate. Dans l’article 5, nous avons établi le protocole d’enregistrement ERG normalisé sur le singe vervet, et nous avons produit un atlas d’ondes ERG spécifique à cette espèce, selon les règles de l’International Society for Clinical Electrophysiology of Vision (ISCEV). Les patrons électrorétinographiques se sont avérés semblables à ceux de l’humain et ont confirmé la similarité entre ces deux espèces. Dans l’article 6, nous avons démontré que le blocage de CB1R ou CB2R entraine une modification de l’électrorétinogramme, tant au niveau photopique que scotopique, ce qui supporte l’implication de ces récepteurs dans la modulation des ondes de l’ERG. Finalement, dans l’article 7, nous avons confirmé le modèle neurochimique proposé dans l’article 3 pour expliquer le rôle fonctionnel de GPR55, en montrant que l’activation ou le blocage de ce récepteur, respectivement par un agoniste (lysophosphatidylglucoside, LPG) ou un antagoniste (CID16020046), entraine soit une augmentation ou une baisse significative de l’ERG scotopique seulement. Ces données, prises ensemble, démontrent que les récepteurs CB1R, CB2R et GPR55 sont exprimés dans des types cellulaires bien distincts de la rétine du singe et ont chacun un rôle spécifique. L’importance de notre travail se manifeste aussi par des applications cliniques en permettant le développement de cibles pharmacologiques potentielles dans le traitement des maladies de la rétine.

Accurate prediction of scorpion toxin functional properties from primary structures

Scorpion toxins are common experimental tools for studies of biochemical and pharmacological properties of ion channels. The number of functionally annotated scorpion toxins is steadily growing, but the number of identified toxin sequences is increasing at much faster pace. With an estimated 100,000 different variants, bioinformatic analysis of scorpion toxins is becoming a necessary tool for their systematic functional analysis. Here, we report a bioinformatics-driven system involving scorpion toxin structural classification, functional annotation, database technology, sequence comparison, nearest neighbour analysis, and decision rules which produces highly accurate predictions of scorpion toxin functional properties. (c) 2005 Elsevier Inc. All rights reserved.

Neuroprotectant effects of iso-osmolar D-mannitol to prevent Pacific ciguatoxin-1 induced alterations in neuronal excitability: A comparison with other osmotic agents and free radical scavengers

The basis for the neuroprotectant effect of D-mannitol in reducing the sensory neurological disturbances seen in ciguatera poisoning, is unclear. Pacific ciguatoxin-1 (P-CTX-1), at a concentration 10 nM, caused a statistically significant swelling of rat sensory dorsal root ganglia (DRG) neurons that was reversed by hyperosmolar 50 MM D-mannitol. However, using electron paramagnetic resonance (EPR) spectroscopy, it was found that P-CTX-1 failed to generate hydroxyl free radicals at concentrations of toxin that caused profound effects on neuronal excitability. Whole-cell patch-clamp recordings from DRG neurons revealed that both hyper- and iso-osmolar 50 MM D-mannitol prevented the membrane depolarisation and repetitive firing of action potentials induced by P-CTX-1. In addition, both hyper- and iso-osmolar 50 MM D-mannitol prevented the hyperpolarising shift in steady-state inactivation and the rise in leakage current through tetrodotoxin (TTX)-sensitive Na-v channels, as well as the increased rate of recovery from inactivation of TTX-resistant Nav channels induced by P-CTX-1. D-Mannitol also reduced, but did not prevent, the inhibition of peak TTX-sensitive and TTX-resistant I-Na amplitude by P-CTX-1. Additional experiments using hyper- and isoosmolar D-sorbitol, hyperosmolar sucrose and the free radical scavenging agents Trolox (R) and L-ascorbic acid showed that these agents, unlike D-mannitol, failed to prevent the effects of P-CTX-1 on spike electrogenesis and Na-v channel gating. These selective actions of D-mannitol indicate that it does not act purely as an osmotic agent to reduce swelling of nerves, but involves a more complex action dependent on the Nav channel subtype, possibly to alter or reduce toxin association. (c) 2005 Elsevier Ltd. All rights reserved.

Conotoxins as research tools and drug leads

The complex mixture of biologically active peptides that constitute the venom of Conus species provides a rich source of ion channel neurotoxins. These peptides, commonly known as conotoxins, exhibit a high degree of selectivity and potency for different ion channels and their subtypes making them invaluable tools for unravelling the secrets of the nervous system. Furthermore, several conotoxin molecules have profound applications in drug discovery, with some examples currently undergoing clinical trials. Despite their relatively easy access by chemical synthesis, rapid access to libraries of conotoxin analogues for use in structure-activity relationship studies still poses a significant limitation. This is exacerbated in conotoxins containing multiple disulfide bonds, which often require synthetic strategies utilising several steps. This review will examine the structure and activity of some of the known classes of conotoxins and will highlight their potential as neuropharmacological tools and as drug leads. Some of the classical and more recent approaches to the chemical synthesis of conotoxins, particularly with respect to the controlled formation of disulfide bonds will be discussed in detail. Finally, some examples of structure-activity relationship studies will be discussed, as well as some novel approaches for designing conotoxin analogues.