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Tetrodotoxin
Kinds of Tetrodotoxin Selected AbstractsA slowly inactivating sodium current (INa2) in the plateau range in canine cardiac Purkinje single cellsEXPERIMENTAL PHYSIOLOGY, Issue 1 2007Mario Vassalle The action potential of Purkinje fibres is markedly shortened by tetrodotoxin, suggesting the possibility that a slowly inactivating sodium current might flow during the plateau. The aim of the present experiments was to investigate, in canine cardiac Purkinje single cells by means of a whole cell patch clamp technique, whether a sodium current slowly inactivates at less negative potentials and (if so) some of its distinctive characteristics. The results showed that a 500 ms depolarizing step from a holding potential of ,90 mV to ,50 mV induced the fast inward current INa (labelled here INa1). With steps to ,40 mV or less negative values, a slowly decaying component (tentatively labelled here INa2) appeared, which peaked at ,30 to ,20 mV and decayed slowly and incompletely during the 500 ms steps. The INa2 was present also during steps to ,10 mV, but then the transient outward current (Ito) appeared. When the holding potential (Vh) was decreased to ,60 to ,50 mV, INa2 disappeared even if a small INa1 might still be present. Tetrodotoxin (30 ,m), lignocaine (100 ,m) and cadmium (0.2 mm; but not manganese, 1 mm) blocked INa2. During fast depolarizing ramps, the rapid inactivation of INa1 was followed by a negative slope region. During repolarizing ramps, a region of positive slope was present, whereas INa1 was absent. At less negative values of Vh, the amplitude of the negative and positive slopes became gradually smaller. Gradually faster ramps increased the magnitude of the negative slope, and tetrodotoxin (30 ,m) reduced or abolished it. Thus, Purkinje cells have a slowly decaying inward current owing to Na+ entry (INa2) that is different in several ways from the fast INa1 and that appears important for the duration of the plateau. [source] Identification of Tetrodotoxin and Fish Species in an Adulterated Dried Mullet Roe Implicated in Food PoisoningJOURNAL OF FOOD SCIENCE, Issue 1 2003Y.W. Hsieh ABSTRACT: There was 1 victim of neurotoxic food poisoning from an adulterated dried mullet roe in Kaohsiung, Taiwan, in March 2001. The victim exhibited typical neurotoxic symptoms. The residue of dried mullet roe retained by the victim was assayed for toxicity and mitochondrial DNA. Its toxicity was 3450 mouse units per gram. The toxin was partially purified and identified as tetrodotoxin and derivative. The sequence of the 376-nucleotide region in the cytochrome b gene of the mitochondrial DNA exhibited the same genotype and the same restriction site for SapI as that of the toxic puffer fish Lagocephalus lunaris. [source] Oxytocin receptor expressed on the smooth muscle mediates the excitatory effect of oxytocin on gastric motility in ratsNEUROGASTROENTEROLOGY & MOTILITY, Issue 4 2009J. Qin Abstract, The aim of this study was to localize oxytocin receptor (OTR) in the stomach and to investigate the effect of OT on gastric motility in rats. Western blot and immunohistochemistry methods were used to localize OTR in stomach. The motility of stomach was recorded in vivo (recording of the intragastric pressure), in vitro (recording of the contraction of muscle strips) and on isolated smooth muscle cells. OTR was expressed on cells of both circular and longitudinal muscle of stomach. Systemic administration of OT induced an early transient decrease and a subsequent increase on intragastric pressure. Devazepide (1 mg kg,1, i.v.), a cholecystokinin-1 (CCK1) receptor antagonist, completely abolished the transient response but did not influence the subsequent one. OT (10,9,10,6 mol L,1) dose-dependently increased the contraction of the muscle strips of gastric body, antrum, and pyloric sphincter, and decreased the average cell length of isolated smooth muscle cells. Tetrodotoxin and atropine did not influence the effect of OT on muscle strips. Pretreatment with atosiban, an OTR antagonist, inhibited the spontaneous contraction of muscle strips and abolished the excitatory effect of OT on the muscle strips and the isolated cells. These results suggest that the OTR is expressed on the smooth muscle of the stomach and mediates excitatory effect of OT on gastric motility. [source] Nitric oxide mediates the inhibitory effect of ethanol on the motility of isolated longitudinal muscle of proximal colon in ratsNEUROGASTROENTEROLOGY & MOTILITY, Issue 6 2007S. L. Wang Abstract, The aim of the present study was to investigate the effect of ethanol on colon motility in rats and to test the possibility that nitric oxide (NO) mediates this effect. Proximal colon longitudinal muscle strips (LM) (8 × 3 mm) cut parallel to the longitudinal muscle fibres of the colon were isolated and mounted in an organ bath. Ethanol (0.57, 0.87 and 1.30 mmol L,1) dose-dependently inhibited the motility of LM. Longitudinal muscle strips from female rats were more sensitive to the inhibitory effect of ethanol than that from male rats. L-NAME (N -nitro- l -arginine methyl ester) (100 ,mol L,1), AG (aminoguanidine) (10 ,mol L,1), ODQ (1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one) (10 ,mol L,1) and PTIO (2-Phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide) (200 ,mol L,1) partly blocked the inhibitory effect of ethanol on LM. Pretreatment with L-NAME, AG, ODQ and PTIO abolished the sex difference of the inhibitory effect of ethanol on LM. Tetrodotoxin (TTX) (10 ,mol L,1) partly blocked the inhibitory effect but did not influence the sex difference. The relaxation of LM induced by SNP (sodium nitroprusside) (0.1,10 ,mol L,1) in female rats was greater than that in male rats. In conclusion, ethanol inhibited the colon motility in vitro. This inhibitory effect on LM was mediated by NO through the iNOS , NO , cGMP pathway. [source] Real-time measurement of serotonin release and motility in guinea pig ileumTHE JOURNAL OF PHYSIOLOGY, Issue 2 2006Paul P. Bertrand Enterochromaffin (EC) cells are sensors that detect chemical or mechanical stimuli and respond with release of serotonin (5-HT). 5-HT activates local motor reflexes, but whether local motor reflexes also evoke 5-HT release is unknown. The aim of the present study was to establish the relationship between the release of 5-HT and the enteric neural circuits controlling the movements of the intestine. Recordings were made from full-thickness preparations of guinea pig ileum using electrochemical techniques with carbon fibre electrodes to measure local concentrations of 5-HT. The tension in the circular muscle (CM) and longitudinal muscle (LM) was recorded with force transducers. The release of 5-HT from the EC cells was detected selectively and the timing of the events quantified. Pressure-evoked peristalsis caused detectable 5-HT release only when the recording site was invaded by a ring of CM contraction. Spontaneous and stretch-evoked reflex contraction of the CM and LM occurred simultaneously with 5-HT release. Paralysis of the smooth muscle significantly reduced the stretch-evoked release. Muscarinic agonists evoked reflexes that were associated with increases in tension in CM and LM simultaneous with 5-HT release. Tetrodotoxin abolished the coordination between the CM contraction and 5-HT release but not the direct activation of the CM and EC cells by the agonists. In conclusion, the correlation between local motor reflexes and 5-HT release observed in the present study is caused primarily by the contraction of the smooth muscle and subsequent deformation of the mucosa. The EC cell is, thus, a site of convergence for mechanical forces that contribute to the release of 5-HT during motor reflexes. [source] Long-lasting contractile action and the inhibitory action of cupric ions on ileal longitudinal muscleAUTONOMIC & AUTACOID PHARMACOLOGY, Issue 4 2004K. Miyazaki Summary 1 Cupric ions (Cu2+), at concentrations above 0.03 mm, induced a progressive increase in the tonic contraction of guinea-pig ileal longitudinal muscle. Maximal contraction of 0.1 mm Cu2+ attained a level above that of the 60-mm K+ -induced tonic response, within 20 min of application. The tension induced by Cu2+ persisted for more than several hours. Tetrodotoxin (3 × 10,6 m) had no effect on the contraction induced by 0.1 mm Cu2+. 2 After incubation in a Ca2+ -free medium, the ileal response to 0.1 mm Cu2+ was lost. Nifedipine, a L-type Ca2+ channel blocker, dose-dependently inhibited contractions induced by Cu2+. 3 As the duration of the first application of 0.1 mm Cu2+ increased above 30 min, after washing with normal medium, the contractile response to a second application of 0.1 mm Cu2+ decreased gradually. After 150 min of the first application of 0.1 mm Cu2+, a second application of Cu2+ could not evoke any contraction. 4 After the application of 0.1 mm Cu2+ for 150 min, when muscles were washed with a medium containing 1 mm EDTA, the response to 0.1 mm Cu2+ returned to a greater extent in the normal Ca2+ medium. 5 In conclusion, Cu2+ (0.1 mm) induced a maximal ileal tension above that of the K-induced tonic response within 20 min. The ileal contraction to Cu2+ persisted for more than several hours and depended on extracellular Ca2+ concentrations. It is possible that a part of Cu2+, bound to a EDTA-inaccessible site, also has a tension inhibitory effect. [source] The regulation of veratridine-stimulated electrogenic ion transport in mouse colon by neuropeptide Y (NPY), Y1 and Y2 receptorsBRITISH JOURNAL OF PHARMACOLOGY, Issue 5 2005Niall P Hyland Neuropeptide Y (NPY) is a prominent enteric neuropeptide with prolonged antisecretory effects in mammalian intestine. Veratridine depolarises neurons consequently causing epithelial anion secretion across mouse colon mucosa. Our aim was to characterise functionally, veratridine-stimulated mucosal responses and to determine the roles for NPY, Y1, and Y2 receptors in modulating these neurogenic effects. Colon mucosae (with intact submucous innervation) from wild-type mice (+/+) and knockouts lacking either NPY (NPY,/,), Y1,/, or Y2,/, were placed in Ussing chambers and voltage clamped at 0 mV. Veratridine-stimulated short-circuit current (Isc) responses in +/+, Y1 or Y2 antagonist pretreated +/+ colon, Y1,/, and NPY,/, colon were insensitive to cholinergic blockade by atropine (At; 1 ,M) and hexamethonium (Hex; 10 ,M). Tetrodotoxin (TTX, 100 nM) abolished veratridine responses, but had no effect upon carbachol (CCh) or vasoactive intestinal polypeptide (VIP)-induced secretory responses. To establish the functional roles for Y1 and Y2 receptors, +/+ tissues were pretreated with either the Y1 or Y2 receptor antagonist (BIBO3304 (300 nM) or BIIE0246 (1 ,M), respectively) and veratridine responses were compared with those from Y1,/, or Y2,/, colon. Neither BIBO3304 nor Y1,/, altered veratridine-induced secretion, but Y1 agonist responses were abolished in both preparations. In contrast, the Y2 antagonist BIIE0246 significantly amplified veratridine responses in +/+ mucosa. Unexpectedly, NPY,/, colon exhibited significantly attenuated veratridine responses (between 1 and 5 min). We demonstrate that electrogenic veratridine responses in mouse colon are noncholinergic and that NPY can act directly upon epithelia, a Y1 receptor effect. The enhanced veratridine response observed in +/+ tissue following BIIE0246, indicates that Y2 receptors are located on submucosal neurons and that their activation by NPY will inhibit enteric noncholinergic secretory neurotransmission. We also demonstrate Y1 and Y2 receptor-mediated antisecretory tone in +/+ colon and show selective loss of each in Y1 and Y2 null colon respectively. In NPY,/, tissue, only Y1 -mediated tone was present, this presumably being mediated by endogenous endocrine peptide YY. Y2 tone was absent from NPY,/, (and Y2,/,) colon and we conclude that NPY activation of neuronal Y2 receptors attenuates secretory neurotransmission thereby providing an absorptive electrolyte tone in isolated colon. British Journal of Pharmacology (2005) 146, 712,722. doi:10.1038/sj.bjp.0706368 [source] Adrenergic mechanisms in canine nasal venous systemsBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2003Min Wang We investigated the adrenergic mechanisms of the two venous systems that drain the nasal mucosa, thereby their exact role in eliciting nasal decongestion. The action of endogenously released noradrenaline and exogenous adrenergic agonists on different segments of the nasal venous systems, i.e. collecting (LCV, SCV) and outflow (SPV) veins of posterior venous system, collecting (ACV) and outflow (DNV) veins of anterior venous system and venous sinusoids of the septal mucosa (SM), were studied. In vitro isometric tension of the vascular segments was measured. Transmural nerve stimulation (TNS) produced constriction in ACV, DNV and SM, primary constriction followed by secondary dilatation in LCV and SCV and dilatation in SPV. Tetrodotoxin (10,6M) abolished all responses. Phentolamine (10,6M), prazosin (10,6M) and rauwolscine (10,7M) inhibited the constriction in all venous vessels. Propranolol (10,6M), atenolol (10,6M) and ICI 118,551 (10,6M) inhibited the relaxation in SPV but not in LCV and SCV. Phenylephrine and clonidine constricted whereas dobutamine and terbutaline relaxed all venous vessels dose-dependently. These results indicate ,1 -, ,2 -, ,1 - and ,2 -adrenoceptors are present in both venous systems. TNS causes constriction of anterior venous system, venous sinusoids and posterior collecting veins primarily via postjunctional ,2 -adrenoceptors but relaxation of posterior outflow vein equally via postjunctional ,1 - and ,2 -adrenoceptors. The combined action of the two adrenergic mechanisms can reduce nasal airway resistance in vivo by decreasing vascular capacitance and enhancing venous drainage via the posterior venous system. British Journal of Pharmacology (2003) 138, 145,155. doi:10.1038/sj.bjp.0705020 [source] Smooth muscle 5-HT2A receptors mediating contraction of porcine isolated proximal stomach stripsBRITISH JOURNAL OF PHARMACOLOGY, Issue 8 2002P Janssen The aim of this study was to characterize the 5-HT receptors involved in the 5-HT-induced contraction of longitudinal muscle (LM) strips of porcine proximal stomach. This was done in a classical organ bath set-up for isotonic measurement. The concentration-contraction curve to 5-HT was not modified by 5-HT3 and 5-HT4 receptor antagonism. Methysergide, ketanserin and mesulergine antagonized the curve to 5-HT. Concomitantly, increasing concentrations of ketanserin and mesulergine progressively revealed a biphasic nature of the 5-HT curve. Ketanserin antagonized the low-affinity receptor while it did not modify the high-affinity receptor. Tetrodotoxin did not influence the concentration-contraction curve to 5-HT neither in the absence nor presence of ketanserin, indicating that nerves are not involved. Ketanserin competitively antagonized the monophasic concentration-response curve to ,-Methyl-5-HT, yielding a Schild slope that was not significantly different from unity. After constraining the Schild slope to unity, a pKB estimate of 8.23±0.90 was obtained. This affinity estimate of ketanserin closely approximates previously reported affinities at 5-HT2A receptors. In the presence of ketanserin (0.1 ,M; exposing the high-affinity receptor), a wide range of 5-HT receptor antagonists covering all 5-HT receptors known, was tested. Only methysergide and ritanserin inhibited the response to 5-HT, thus expressing affinity for the high-affinity receptor. This did not reveal the identity of the receptor involved. It can be concluded that 5-HT induces pig proximal stomach (LM) contraction via 5-HT2A receptors located on smooth muscle. A ketanserin-insensitive phase of contractions could not be characterized between the actually known classes of 5-HT receptors with the pharmacological tools that were used. British Journal of Pharmacology (2002) 137, 1217,1224. doi:10.1038/sj.bjp.0704992 [source] Regional variation in electrically-evoked contractions of rabbit isolated pulmonary arteryBRITISH JOURNAL OF PHARMACOLOGY, Issue 4 2002V Margaret Jackson Electrically-evoked contractions in different regions of the rabbit isolated pulmonary artery have been investigated using stimulation parameters generally assumed to stimulate nerves selectively. In extrapulmonary artery, trains of stimuli (10 Hz; pulse width 0.1 ms) evoked monophasic contractions. In contrast, a biphasic contraction was evoked in the intrapulmonary artery consisting of an initial fast component followed by a secondary very long-lasting component. The contraction in the extrapulmonary artery was prazosin-sensitive (1 ,M) whereas that in the intrapulmonary artery was prazosin-resistant. ,,,-Methylene ATP (1 ,M), atropine (1 ,M), losartan (1 ,M), BIBO3304 (1 nM) or nifedipine (1 ,M) had no effect on the biphasic contraction of the intrapulmonary artery. Bretylium (2 ,M) abolished the contraction of extrapulmonary artery but only partially inhibited the initial component in the intra region with no effect on the second component. Tetrodotoxin (0.3,1 ,M), abolished the contraction of extrapulmonary artery but only partially reduced the electrically-evoked contraction of intrapulmonary artery. Removal of the endothelium and application of sulphisoxazole (0.6,22 ,M) had no effect. Varying the resting tone on the arteries, or applying gadolinium, had no effect on contractions. Using confocal microscopy and calcium imaging, reproducible whole cell calcium transients were evoked in individual smooth muscle cells in intact preparations but only when direct muscle stimulation was used (pulse width of 5,10 ms). No detectable changes in calcium were elicited when brief pulse widths were used (0.1,2 ms). Together, these data suggest that noradrenaline is the neurotransmitter inducing contraction in extrapulmonary artery. Noradrenaline and sympathetic nerves appear to play a less important role in the intrapulmonary artery. The tetrodoxin-resistant component is not mediated by ATP, NPY, acetylcholine, angiotensins, ET-1, stretch-activation or Ca2+ influx through L-type Ca2+ channels. Smooth muscle cells do not appear to be damaged by the stimulation protocol. The mechanism underlying the long lasting contraction of intrapulmonary artery evoked by brief electrical stimuli remains to be elucidated. British Journal of Pharmacology (2002) 137, 488,496. doi:10.1038/sj.bjp.0704863 [source] Biphasic effects of NMDA on the motility of the rat portal veinBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2000Z L Rossetti The effect of NMDA on the motility of the rat portal vein was studied in an isolated preparation. NMDA induced a concentration-dependent (10,7,10,4 M) increase of the contraction frequency (maximum increase, 148±6% of control at NMDA 10,4 M). The NMDA-induced excitatory response was prevented by the competitive NMDA receptor antagonists (±)-2-Amino-5-phosphonopentanoic acid (AP-5, 5×10,4 M) or (RS)-3-(2-carboxypiperazine-4-yl) propyl-1-phosphonic acid (CPP, 10,4 M). Tetrodotoxin (TTX, 10,6 M) or atropine (10,4 M) abolished the NMDA-induced increase of the portal vein motility and reversed the excitatory effect to a concentration-dependent inhibition (maximum inhibition, 52±8 and 29±7% of controls, respectively, at NMDA 10,3 M). Removal of the endothelium abolished the NMDA-induced inhibitory response. Sodium nitroprusside concentration-dependently (10,7,10,5 M) inhibited the portal vein motility, while L -NG -nitro-arginine methyl ester (L -NAME, 10,4 M) reversed the inhibitory effect of NMDA (in the presence of TTX), restoring the portal vein spontaneous activity to control values. These results show that NMDA modulates the portal vein motility in a biphasic manner: via indirect activation, through prejunctional NMDA receptors presumably located on intrinsic excitatory neuronal afferences, or via direct inhibition, through endothelial NMDA receptors activating the nitric oxide pathway. Overall these findings support the hypothesis of the existence of a peripheral glutamatergic innervation modulating the contractile activity of the rat portal vein. British Journal of Pharmacology (2000) 129, 156,162; doi:10.1038/sj.bjp.0703002 [source] An Efficient Total Synthesis of Optically Active Tetrodotoxin,from,LevoglucosenoneCHEMISTRY - AN ASIAN JOURNAL, Issue 1-2 2006Daisuke Urabe Abstract Tetrodotoxin, a toxic principal of puffer-fish poisoning, is one of the most famous marine natural products, and has been known as a formidable synthetic target in synthesis owing to its multifunctional structure and unusual chemical properties. From the perspective of supplying tetrodotoxin derivatives such as labeled molecules for biochemical research, we have completed our second total synthesis of tetrodotoxin from a synthetic intermediate for 11-deoxytetrodotoxin, which was previously prepared from levoglucosenone as a chiral starting material in this laboratory. This paper discloses the details of the total synthesis with special reference to significant influences on the neighboring functional groups found in the installation of guanidine. The established route should allow us to prepare the tetrodotoxin-related compounds required for biochemical studies. [source] Cardiac basal metabolism: energetic cost of calcium withdrawal in the adult rat heartACTA PHYSIOLOGICA, Issue 3 2010P. Bonazzola Abstract Aim:, Cardiac basal metabolism upon extracellular calcium removal and its relationship with intracellular sodium and calcium homeostasis was evaluated. Methods:, A mechano-calorimetric technique was used that allowed the simultaneous and continuous measurement of both heat rate and resting pressure in arterially perfused quiescent adult rat hearts. Using pharmacological tools, the possible underlying mechanisms related to sodium and calcium movements were investigated. Results:, Resting heat rate (expressed in mW g,1dry wt) increased upon calcium withdrawal (+4.4 ± 0.2). This response was: (1) unaffected by the presence of tetrodotoxin (+4.3 ± 0.6), (2) fully blocked by both, the decrease in extracellular sodium concentration and the increase in extracellular magnesium concentration, (3) partially blocked by the presence of either nifedipine (+2.8 ± 0.4), KB-R7943 (KBR; +2.5 ± 0.2), clonazepam (CLO; +3.1 ± 0.3) or EGTA (+1.9 ± 0.3). The steady heat rate under Ca2+ -free conditions was partially reduced by the addition of Ru360 (,1.1 ± 0.2) but not CLO in the presence of EGTA, KBR or Ru360. Conclusion:, Energy expenditure for resting state maintenance upon calcium withdrawal depends on the intracellular rise in both sodium and calcium. Our data are consistent with a mitochondrial Ca2+ cycling, not detectable under normal calcium diastolic levels. The experimental condition here analysed, partially simulates findings reported under certain pathological situations including heart failure in which mildly increased levels of both diastolic sodium and calcium have also been found. Therefore, under such pathological conditions, hearts should distract chemical energy to fuel processes associated with sodium and calcium handling, making more expensive the maintenance of their functions. [source] Three-dimensional slice cultures from murine fetal gut for investigations of the enteric nervous systemDEVELOPMENTAL DYNAMICS, Issue 1 2007Marco Metzger Abstract Three-dimensional intestinal cultures offer new possibilities for the examination of growth potential, analysis of time specific gene expression, and spatial cellular arrangement of enteric nervous system in an organotypical environment. We present an easy to produce in vitro model of the enteric nervous system for analysis and manipulation of cellular differentiation processes. Slice cultures of murine fetal colon were cultured on membrane inserts for up to 2 weeks without loss of autonomous contractility. After slice preparation, cultured tissue reorganized within the first days in vitro. Afterward, the culture possessed more than 35 cell layers, including high prismatic epithelial cells, smooth muscle cells, glial cells, and neurons analyzed by immunohistochemistry. The contraction frequency of intestinal slice culture could be modulated by the neurotransmitter serotonin and the sodium channel blocker tetrodotoxin. Coculture experiments with cultured neurospheres isolated from enhanced green fluorescent protein (eGFP) transgenic mice demonstrated that differentiating eGFP-positive neurons were integrated into the intestinal tissue culture. This slice culture model of enteric nervous system proved to be useful for studying cell,cell interactions, cellular signaling, and cell differentiation processes in a three-dimensional cell arrangement. Developmental Dynamics 236:128,133, 2007. © 2006 Wiley-Liss, Inc. [source] Activation of a calcium entry pathway by sodium pyrithione in the bag cell neurons of AplysiaDEVELOPMENTAL NEUROBIOLOGY, Issue 4 2004Ronald J. Knox Abstract The ability of sodium pyrithione (NaP), an agent that produces delayed neuropathy in some species, to alter neuronal physiology was accessed using ratiometric imaging of cytosolic free Ca2+ concentration ([Ca2+]i) in fura PE-filled cultured Aplysia bag cell neurons. Bath-application of NaP evoked a [Ca2+]i elevation in both somata and neurites with an EC50 of ,300 nM and a Hill coefficient of ,1. The response required the presence of external Ca2+, had an onset of 3,5 min, and generally reached a maximum within 30 min. 2-Methyl-sulfonylpyridine, a metabolite and close structural analog of NaP, did not elevate [Ca2+]i. Under whole-cell current-clamp recording, NaP produced a ,14 mV depolarization of resting membrane potential that was dependent on external Ca2+. These data suggested that NaP stimulates Ca2+ entry across the plasma membrane. To minimize the possibility that a change in cytosolic pH was the basis for NaP-induced Ca2+ entry, bag cell neuron intracellular pH was estimated with the dye 2,,7,-bis(carboxyethyl-5(6)-carboxy-fluorescein acetoxy methylester. Exposure of the neurons to NaP did not alter intracellular pH. The slow onset and sustained nature of the NaP response suggested that a cation exchange mechanism coupled either directly or indirectly to Ca2+ entry could underlie the phenomenon. However, neither ouabain, a Na+/K+ ATPase inhibitor, nor removal of extracellular Na+, which eliminates Na+/Ca2+ exchanger activity, altered the NaP-induced [Ca2+]i elevation. Finally, the possibility that NaP gates a Ca2+ -permeable ion channel in the plasma membrane was examined. NaP did not appear to activate two major forms of bag cell neuron Ca2+ -permeable ion channels, as Ca2+ entry was unaffected by inhibition of voltage-gated Ca2+ channels using nifedipine or by inhibition of a voltage-dependent, nonselective cation channel using a high concentration of tetrodotoxin. In contrast, two potential store-operated Ca2+ entry current inhibitors, SKF-96365 and Ni2+, attenuated NaP-induced Ca2+ entry. We conclude that NaP activates a slow, persistent Ca2+ influx in Aplysia bag cell neurons. © 2004 Wiley Periodicals, Inc. J Neurobiol 411,423, 2004 [source] Nicotinic synapses formed between chick ciliary ganglion neurons in culture resemble those present on the neurons in vivoDEVELOPMENTAL NEUROBIOLOGY, Issue 4 2001Min Chen Abstract We studied nicotinic synapses between chick ciliary ganglion neurons in culture to learn more about factors influencing their formation and receptor subtype dependence. After 4,8 days in culture, nearly all neurons displayed spontaneous excitatory postsynaptic currents (sEPSCs), which occurred at about 1 Hz. Neurons treated with tetrodotoxin displayed miniature EPSCs (mEPSCs), but these occurred at low frequency (0.1 Hz), indicating that most sEPSCs are actually impulse driven. The sEPSCs could be classified by decay kinetics as fast, slow, or biexponential and, reminiscent of the situation in vivo, were mediated by two major nicotinic acetylcholine receptor (AChR) subtypes. Fast sEPSCs were blocked by ,-bungarotoxin (,Bgt), indicating dependence on ,Bgt-AChRs, most of which are ,7 subunit homopentamers. Slow sEPSCs were unaffected by ,Bgt, and were blocked instead by the ,3/,2-selective ,-conotoxin-MII (,CTx-MII), indicating dependence on ,3*-AChRs, which lack ,7 and contain ,3 subunits. Biexponential sEPSCs were mediated by both ,Bgt- and ,3*-AChRs because they had fast and slow components qualitatively similar to those comprising simple events, and these were reduced by ,Bgt and blocked by ,CTx-MII, respectively. Fluorescence labeling experiments revealed both ,Bgt- and ,3*-AChR clusters on neuron somata and neurites. Colabeling with antisynaptic vesicle protein antibody suggested that some ,3*-AChR clusters, and a few ,Bgt-AChR clusters are associated with synaptic sites, as is the case in vivo. These findings demonstrate the utility of ciliary ganglion neuron cultures for studying the regulation of nicotinic synapses, and suggest that mixed AChR subtype synapses characteristic of the neurons in vivo can form in the absence of normal inputs or targets. © 2001 John Wiley & Sons, Inc. J Neurobiol 47: 265,279, 2001 [source] A new animal model of infantile spasms with unprovoked persistent seizuresEPILEPSIA, Issue 2 2008Chong L. Lee Summary Purpose: Infantile spasms is one of the most severe epileptic syndromes of infancy and early childhood. Progress toward understanding the pathophysiology of this disorder and the development of effective therapies has been hindered by the lack of a relevant animal model. We report here the creation of such a model. Methods: The sodium channel blocker, tetrodotoxin (TTX), was chronically infused into the developing neocortex or hippocampus of infant rats by way of an osmotic minipump starting on postnatal day 10,12. Results: After a minimum of 10 days of infusion, approximately one-third of these rats began to display very brief (1,2 s) spasms, which consisted of symmetric or asymmetric flexion or extension of the trunk and sometimes involvement of one or both forelimbs. The typical ictal EEG pattern associated with the behavioral spasms consisted of an initial generalized, high amplitude, slow wave followed by an electrodecrement with superimposed fast activity. The interictal EEG revealed multifocal spikes and sharp waves, and in most animals that had spasms a hypsarrhythmic pattern was seen, at least intermittently, during NREM sleep. Like in humans, the spasms in the rat often occurred in clusters especially during sleep,wake transitions. Comparison of the ictal and interictal EEGs recorded in this model and those from humans with infantile spasms revealed that the patterns and the frequency components of both the ictal events and hypsarrhythmia were very similar. Discussion: The TTX model of infantile spasms should be of value in furthering an understanding of the pathophysiology of this seizure disorder. [source] Abnormal Excitability of Hippocampal CA3 Neurons in Noda Epileptic Rat (NER): Alteration of Seizure with AgingEPILEPSIA, Issue 2000Ryosuke Hanaya Purpose: Noda epileptic rat (NER), a mutant found in thc colony of Crj:Wistar rats, spontaneously shows tonic-clonic convulsions approximately once every 30 hours from 8,16 weeks of age. A long-lasting dcpolarization shift accompanied by repetitivc firings are observed in hippocampal CA3 pyramidal neurons of NER with seizures. Using hippocampal slice preparations of NER, the present electrophysiologi- cal study was performed to elucidate whether this abnormal firing in CA3 neurons developed with age and if abnormality of Ca2+ channel was involved. Methods: Hippocampal slices (40Opm) werc prepared from NER and normal Wistar rats (age; 4,29 weeks). A single rectangular pulse stimulus composed of 0.1-ms duration was delivered to the mossy fibers every 5 seconds though a bipolar electrode placed in the granular cell layer of the dentate gyrus. Intracellular recording was made from the CA3 pyramidal cell using a microelectrode containing 3M KCI intracellular recordings. A Ca2+ spike was elicited by applying a depolarizing pulse (InA, 120ms) in the cell through the recording electrode under a blockadc of Na+ and K+ channels using 1 pM tetrodotoxin and I 0mM tctraethylammonium added to the artificial CSF, respectivcly. Nicardipine (I-IOOnM), a Ca2+ channel blocker, was applicd to the bath. Results: Thirty-seven slices from I9 NER and 6 slices from 4 normal Wishe rats were used. There were no obvious changes in the resting membrane potentials of CA3 neurons between NER and Wistar rats tested. When a single stimulus was delivered to the mossy fibers, a long-lasting depolarization shift accompanied by repetitive firings followed by after-hyperpolarization werc also obtained i n hippocampal CA3 neurons of young NER (4,5 weeks of age) before occurrence of any seizurcs, although the depolarization shift in younger NER was shorter than that in NER aged more than 6 weeks. These abnormal firings werc evokcd in 58% and 30% of all CA3 neurons tested in the younger and mature NER (6,1 5 weeks of age), respectively. Furthermore, abnormal firing was not elicited in NER aged after I6 weeks. Agc-matched Wistar rats showed only single action potentials without any depolarization shift with single mossy fiber stimulation. Bath application of nicardipine (IOnM) inhibited this long-lasting depolarization shift and the accompanying repetitive firing followed by afterhypcrpolarization without affecting the first spike induced by mossy fiber stimulations. Furthermore, nicai-dipine (IOnM) inhibited the Ca2+ spikes elicited by applying a depolarizing pulse in the neurons of NER with seizures, although a higher dose (100nM) did not affect those in Wistar rats. Conclusions: These findings indicate that abnormal excitability of the NER CA3 pyramidal neurons is probably due to abnormality in the Ca2+ channcls. The abnorinal excitability was observed in NER at an age when tonic-clonic convulsions were not detected, suggesting that thc hippocampus may probably scrve as an epileptogenic focus in younger NER and the seizure impulses originating i n this area are transinittcd to the new other seizurc foci in mature NER. [source] Callosal contribution to ocular dominance in rat primary visual cortexEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 7 2010Chiara Cerri Abstract Ocular dominance (OD) plasticity triggered by monocular eyelid suture is a classic paradigm for studying experience-dependent changes in neural connectivity. Recently, rodents have become the most popular model for studies of OD plasticity. It is therefore important to determine how OD is determined in the rodent primary visual cortex. In particular, cortical cells receive considerable inputs from the contralateral hemisphere via callosal axons, but the role of these connections in controlling eye preference remains controversial. Here we have examined the role of callosal connections in binocularity of the visual cortex in naïve young rats. We recorded cortical responses evoked by stimulation of each eye before and after acute silencing, via stereotaxic tetrodotoxin (TTX) injection, of the lateral geniculate nucleus ipsilateral to the recording site. This protocol allowed us to isolate visual responses transmitted via the corpus callosum. Cortical binocularity was assessed by visual evoked potential (VEP) and single-unit recordings. We found that acute silencing of afferent geniculocortical input produced a very significant reduction in the contralateral-to-ipsilateral (C/I) VEP ratio, and a marked shift towards the ipsilateral eye in the OD distribution of cortical cells. Analysis of absolute strength of each eye indicated a dramatic decrease in contralateral eye responses following TTX, while those of the ipsilateral eye were reduced but maintained a more evident input. We conclude that callosal connections contribute to normal OD mainly by carrying visual input from the ipsilateral eye. These data have important implications for the interpretation of OD plasticity following alterations of visual experience. [source] A novel role for MNTB neuron dendrites in regulating action potential amplitude and cell excitability during repetitive firingEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 12 2008Richardson N. Leão Abstract Principal cells of the medial nucleus of the trapezoid body (MNTB) are simple round neurons that receive a large excitatory synapse (the calyx of Held) and many small inhibitory synapses on the soma. Strangely, these neurons also possess one or two short tufted dendrites, whose function is unknown. Here we assess the role of these MNTB cell dendrites using patch-clamp recordings, imaging and immunohistochemistry techniques. Using outside-out patches and immunohistochemistry, we demonstrate the presence of dendritic Na+ channels. Current-clamp recordings show that tetrodotoxin applied onto dendrites impairs action potential (AP) firing. Using Na+ imaging, we show that the dendrite may serve to maintain AP amplitudes during high-frequency firing, as Na+ clearance in dendritic compartments is faster than axonal compartments. Prolonged high-frequency firing can diminish Na+ gradients in the axon while the dendritic gradient remains closer to resting conditions; therefore, the dendrite can provide additional inward current during prolonged firing. Using electron microscopy, we demonstrate that there are small excitatory synaptic boutons on dendrites. Multi-compartment MNTB cell simulations show that, with an active dendrite, dendritic excitatory postsynaptic currents (EPSCs) elicit delayed APs compared with calyceal EPSCs. Together with high- and low-threshold voltage-gated K+ currents, we suggest that the function of the MNTB dendrite is to improve high-fidelity firing, and our modelling results indicate that an active dendrite could contribute to a ,dual' firing mode for MNTB cells (an instantaneous response to calyceal inputs and a delayed response to non-calyceal dendritic excitatory postsynaptic potentials). [source] GAT-1 acts to limit a tonic GABAA current in rat CA3 pyramidal neurons at birthEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 3 2007Sampsa T. Sipilä Abstract Tonic activation of GABAA receptors takes place before the development of functional synapses in cortical structures. We studied whether inefficient GABA uptake might explain the presence of a tonic GABAA -mediated current (IGABA-A) in early postnatal hippocampal pyramidal neurons. The data show, however, that the tonic IGABA-A is enhanced by the specific blocker of GABA transporter-1 (GAT-1), NO-711 (1-[2-[[(Diphenylmethyleneimino]oxy]ethyl]-1,2,5,6-tetrahydro-3-pyridinecarboxylic acid hydrochloride), at birth in rat CA3 pyramidal neurons. NO-711 also prolonged the duration of GABA transients during endogenous hippocampal network events (known as giant depolarizing potentials) at postnatal day 0. The endogenous tonic IGABA-A was seen and it was enhanced by NO-711 in the presence of tetrodotoxin, which itself had only a minor effect on the holding current under control conditions. This indicates that the source of interstitial GABA is largely independent of action-potential activity. The tonic IGABA-A in neonatal CA3 pyramidal neurons was increased by zolpidem, indicating that at least a proportion of the underlying GABAA receptors contain ,2 and ,1,,3 subunits. The present data point to a significant role for GAT-1 in the control of the excitability of immature hippocampal neurons and networks. [source] Prokineticin 2 depolarizes paraventricular nucleus magnocellular and parvocellular neuronsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 2 2007Erik A. Yuill Abstract Blind whole-cell patch-clamp techniques were used to examine the effects of prokineticin 2 (PK2) on the excitability of magnocellular (MNC), parvocellular preautonomic (PA), and parvocellular neuroendocrine (NE) neurons within the hypothalamic paraventricular nucleus (PVN) of the rat. The majority of MNC neurons (76%) depolarized in response to 10 nm PK2, effects that were eliminated in the presence of tetrodotoxin (TTX). PK2 also caused an increase in excitatory postsynaptic potential (EPSP) frequency, a finding that was confirmed by voltage clamp recordings demonstrating increases in excitatory postsynaptic current (EPSC) frequency. The depolarizing effects of PK2 on MNC neurons were also abolished by kynurenic acid (KA), supporting the conclusion that the effects of PK2 are mediated by the activation of glutamate interneurons within the hypothalamic slice. PA (68%) and NE (67%) parvocellular neurons also depolarized in response to 10 nm PK2. However, in contrast to MNC neurons, these effects were maintained in TTX, indicating that PK2 directly affects PA and NE neurons. PK2-induced depolarizations observed in PA and NE neurons were found to be concentration-related and receptor mediated, as experiments performed in the presence of A1MPK1 (a PK2 receptor antagonist) abolished the effects of PK2 on these subpopulations of neurons. The depolarizing effects of PK2 on PA and NE neurons were also shown to be abolished by PD 98059 (a mitogen activated protein kinase (MAPK) inhibitor) suggesting that PK2 depolarizes PVN parvocellular neurons through a MAPK signalling mechanism. In combination, these studies have identified separate cellular mechanisms through which PK2 influences the excitability of different subpopulations of PVN neurons. [source] Disparate cholinergic currents in rat principal trigeminal sensory nucleus neurons mediated by M1 and M2 receptors: a possible mechanism for selective gating of afferent sensory neurotransmissionEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 12 2006Kristi A. Kohlmeier Abstract Neurons situated in the principal sensory trigeminal nucleus (PSTN) convey orofacial sensory inputs to thalamic relay regions and higher brain centres, and the excitability of these ascending tract cells is modulated across sleep/wakefulness states and during pain conditions. Moreover, acetylcholine release changes profoundly across sleep/wakefulness states and ascending sensory neurotransmission is altered by cholinergic agonists. An intriguing possibility is, therefore, that cholinergic mechanisms mediate such state-dependent modulation of PSTN tract neurons. We tested the hypotheses that cholinergic agonists can modulate PSTN cell excitability and that such effects are mediated by muscarinic receptor subtypes, using patch-clamp methods in rat and mouse. In all examined cells, carbachol elicited an electrophysiological response that was independent of action potential generation as it persisted in the presence of tetrodotoxin. Responses were of three types: depolarization, hyperpolarization or a biphasic response consisting of hyperpolarization followed by depolarization. In voltage-clamp mode, carbachol evoked corresponding inward, outward or biphasic currents. Moreover, immunostaining for the vesicle-associated choline transporter showed cholinergic innervation of the PSTN. Using muscarinic receptor antagonists, we found that carbachol-elicited PSTN neuron hyperpolarization was mediated by M2 receptors and depolarization, in large part, by M1 receptors. These data suggest that acetylcholine acting on M1 and M2 receptors may contribute to selective excitability enhancement or depression in individual, rostrally projecting sensory neurons. Such selective gating effects via cholinergic input may play a functional role in modulation of ascending sensory transmission, including across behavioral states typified by distinct cholinergic tone, e.g. sleep/wakefulness arousal levels or neuropathic pain conditions. [source] Differential responses to NMDA receptor activation in rat hippocampal interneurons and pyramidal cells may underlie enhanced pyramidal cell vulnerabilityEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 12 2005E. Avignone Abstract Hippocampal interneurons are generally more resistant than pyramidal cells to excitotoxic insults. Because NMDA receptors play a crucial role in neurodegeneration, we have compared the response to exogenous NMDA in CA1 pyramidal cells and interneurons of the stratum oriens using combined whole-cell patch-clamp recording and ratiometric Ca2+ imaging. In voltage-clamp, current-clamp or in nominally Mg2+ -free medium, NMDA (10 µm; 3,5 min exposure in the presence of tetrodotoxin) induced a markedly larger inward current and Ca2+ rise in pyramidal cells than in interneurons. Pyramidal cells also showed a more pronounced voltage dependence in their response to NMDA. We hypothesized that this enhanced response to NMDA receptor activation in pyramidal cells could underlie their increased vulnerability to excitotoxicity. Using loss of dye as an indicator of degenerative membrane disruption, interneurons tolerated continuous exposure to a high concentration of NMDA (30 µm) for longer periods than pyramidal cells. This acute neurodegeneration in pyramidal cells was independent of intracellular Ca2+, because high intracellular BAPTA (20 mm) did not prolong survival time. Thus, a plausible explanation for the enhanced sensitivity of pyramidal neurons to excitotoxic insults associated with cerebral ischemia is their greater response to NMDA receptor activation, which may reflect differences in NMDA receptor expression and/or subunit composition. [source] Activity-dependent subcellular localization of NAC1EUROPEAN JOURNAL OF NEUROSCIENCE, Issue 2 2005Laxman Korutla Abstract The expression of the transcriptional regulator NAC1 is increased in the nucleus accumbens of rats withdrawn from cocaine self-administration, and in vivo studies indicate that the up-regulation is a compensatory mechanism opposing the acute effects of cocaine. Both mammalian two-hybrid assay and punctate localization largely in the nucleus suggest NAC1 is a transcriptional regulator. However, in this report it is shown that in differentiated PC12 and Neuro2A cells, as well as in primary cortical neurons, NAC1 is diffusely expressed not only in the cell nucleus but also in cytoplasm. Blockade of spontaneous electrical activity by tetrodotoxin prevented the diffuse expression of NAC1, and depolarization with high potassium concentrations induced diffuse cellular localization in non-differentiating cells. The use of protein kinase C (PKC) inhibitors and activator, as well as the systematic mutation of potential PKC phosphorylation sites in NAC1, demonstrated that phosphorylation of residue S245 by PKC is a necessary event inducing diffuse NAC1 expression outside of the nucleus. These observations indicate a potential non-transcriptional role for NAC1 in the brain. [source] Distribution and functional characterization of human Nav1.3 splice variantsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2005R. Thimmapaya Abstract The focus of the present study is the molecular and functional characterization of four splice variants of the human Nav1.3 , subunit. These subtypes arise due to the use of alternative splice donor sites of exon 12, which encodes a region of the , subunit that resides in the intracellular loop between domains I and II. This region contains several important phosphorylation sites that modulate Na+ channel kinetics in related sodium channels, i.e. Nav1.2. While three of the four Nav1.3 isoforms, 12v1, 12v3 and 12v4 have been previously identified in human, 12v2 has only been reported in rat. Herein, we evaluate the distribution of these splice variants in human tissues and the functional characterization of each of these subtypes. We demonstrate by reverse transcriptase-polymerase chain reaction (RT-PCR) that each subtype is expressed in the spinal cord, thalamus, amygdala, cerebellum, adult and fetal whole brain and heart. To investigate the functional properties of these different splice variants, each , subunit isoform was cloned by RT-PCR from human fetal brain and expressed in Xenopus oocytes. Each isoform exhibited functional voltage-dependent Na+ channels with similar sensitivities to tetrodotoxin (TTX) and comparable current amplitudes. Subtle shifts in the V1/2 of activation and inactivation (2,3 mV) were observed among the four isoforms, although the functional significance of these differences remains unclear. This study has demonstrated that all four human splice variants of the Nav1.3 channel , subunit are widely expressed and generate functional TTX-sensitive Na+ channels that likely modulate cellular excitability. [source] Activity-dependent maturation of excitatory synaptic connections in solitary neuron cultures of mouse neocortexEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 2 2005Naoki Takada Abstract Activity plays important roles in the formation and maturation of synaptic connections. We examined these roles using solitary neocortical excitatory neurons, receiving only self-generated synaptic inputs, cultured in a microisland with and without spontaneous spike activity. The amplitude of excitatory postsynaptic currents (EPSCs), evoked by applying brief depolarizing voltage pulses to the cell soma, continued to increase from 7 to 14 days in culture. Short-term depression of EPSCs in response to paired-pulse or 10-train-pulse stimulation decreased with time in culture. These developmental changes were prevented when neurons were cultured in a solution containing tetrodotoxin (TTX). The number of functional synapses estimated by recycled synaptic vesicles with FM4-64 was significantly smaller in TTX-treated than control neurons. However, the miniature EPSC amplitude remained unchanged during development, irrespective of activity. Transmitter release probability, assessed by use-dependent blockade of N -methyl- d -aspartate receptor-mediated EPSCs with MK-801, was higher in TTX-treated than control neurons. Therefore, the activity-dependent increase in EPSC amplitude was mainly ascribed to the increase in synapse number, while activity-dependent alleviation of short-term depression was mostly ascribed to the decrease in release probability. The effect of activity blockade on short-term depression, but not EPSC amplitude, was reversed after 4 days of TTX removal, indicating that synapse number and release probability are controlled by activity in very different ways. These results demonstrate that activity regulates the conversion of immature synapses transmitting low-frequency input signals preferentially to mature synapses transmitting both low- and high-frequency signals effectively, which may be necessary for information processing in mature cortex. [source] Electrical and neurotransmitter activity of mature neurons derived from mouse embryonic stem cells by Sox-1 lineage selection and directed differentiationEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 12 2004R. J. Lang Abstract Sx1TV2/16C is a mouse embryonic stem (ES) cell line in which one copy of the Sox1 gene, an early neuroectodermal marker, has been targeted with a neomycin (G418) selection cassette. A combination of directed differentiation with retinoic acid and G418 selection results in an enriched neural stem cell population that can be further differentiated into neurons. After 6,7 days post-plating (D6,7PP) most neurons readily fired tetrodotoxin (TTX)-sensitive action potentials due to the expression of TTX-sensitive Na+ and tetraethylammonium (TEA)-sensitive K+ channels. Neurons reached their maximal cell capacitance after D6,7PP; however, ion channel expression continued until at least D21PP. The percentage of cells receiving spontaneous synaptic currents (s.s.c.) increased with days in culture until 100% of cells received a synaptic input by D20PP. Spontaneous synaptic currents were reduced in amplitude and frequency by TTX, or upon exposure to a Ca2+ -free, 2.5 mm Mg2+ saline. S.s.c. of rapid decay time constants were preferentially blocked by the nonNMDA glutamatergic receptor antagonists CNQX or NBQX. Ca2+ levels within ES cell-derived neurons increased in response to glutamate receptor agonists l -glutamate, AMPA, N -methyl- d -aspartate (NMDA) and kainic acid and to acetylcholine, ATP and dopamine. ES cell-derived neurons also generated cationic and Cl, -selective currents in response to NMDA and glycine or GABA, respectively. It was concluded that ES-derived neurons fire action potentials, receive excitatory and inhibitory synaptic input and respond to various neurotransmitters in a manner akin to primary central neurons. [source] Synaptic and non-synaptic mechanisms of amygdala recruitment into temporolimbic epileptiform activitiesEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 10 2003Julia Klueva Abstract Lateral amygdala (LA) activity during synchronized-epileptiform discharges in temporolimbic circuits was investigated in rat horizontal slices containing the amygdala, hippocampus (Hip), perirhinal (Prh) and lateral entorhinal (LEnt) cortex, through multiple-site extra- and intracellular recording techniques and measurement of the extracellular K+ concentration. Application of 4-aminopyridine (50 µm) induced epileptiform discharges in all regions under study. Slow interictal-like burst discharges persisted in the Prh/LEnt/LA after disconnection of the Hip, seemed to originate in the Prh as shown from time delay analyses, and often preceded the onset of ictal-like activity. Disconnection of the amygdala resulted in de-synchronization of epileptiform discharges in the LA from those in the Prh/LEnt. Interictal-like activity was intracellularly reflected in LA projection neurons as ,-aminobutyric acid (GABA)A/B receptor-mediated synaptic responses, and depolarizing electrogenic events (spikelets) residing on the initial phase of the GABA response. Spikelets were considered antidromically conducted ectopic action potentials generated at axon terminals, as they were graded in amplitude, were not abolished through hyperpolarizing membrane responses (which effectively blocked evoked orthodromic action potentials), lacked a clear prepotential or synaptic potential, were not affected through blockers of gap junctions, and were blocked through remote application of tetrodotoxin at putative target areas of LA projection neurons. Remote application of a GABAB receptor antagonist facilitated spikelet generation. A transient elevation in the extracellular K+ level averaging 3 mm above baseline occurred in conjunction with interictal-like activity in all areas under study. We conclude that interictal-like discharges in the LA/LEnt/Prh spread in a predictable manner through the synaptic network with the Prh playing a leading role. The rise in extracellular K+ may provide a depolarizing mechanism for recruitment of interneurons and generation of ectopic action potentials at axon terminals of LA projection neurons. Antidromically conducted ectopic action potentials may provide a spreading mechanism of seizure activity mediated by diffuse axonal projections of LA neurons. [source] Ionic currents underlying rhythmic bursting of ventral mossy cells in the developing mouse dentate gyrusEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 7 2003Shozo Jinno Abstract The electrophysiological properties of mossy cells were examined in developing mouse hippocampal slices using whole-cell patch-clamp techniques, with particular reference to the dorsoventral difference. Dorsal mossy cells exhibited a higher spontaneous excitatory postsynaptic potential (EPSP) frequency and larger maximal EPSP amplitude than ventral mossy cells. On the other hand, the blockade of synaptic inputs with glutamatergic and GABAergic antagonists disclosed a remarkable dorsoventral difference in the intrinsic activity: none (0/27) of the dorsal mossy cells showed intrinsic bursting, whereas the majority (35/47) of the ventral mossy cells exhibited intrinsic rhythmic bursting. To characterize the ionic currents underlying the rhythmic bursting of mossy cells, we used somatic voltage-clamp recordings in the subthreshold voltage range. Ventral bursting cells possessed both hyperpolarization-activated current (Ih) and persistent sodium current (INaP), whereas dorsal and ventral nonbursting cells possessed Ih but no INaP. Blockade of Ih with cesium did not affect the intrinsic bursting of ventral mossy cells. In contrast, the blockade of INaP with tetrodotoxin or phenytoin established a stable subthreshold membrane potential in ventral bursting cells. The current,voltage curve of ventral bursting cells showed a region of tetrodotoxin-sensitive negative slope conductance between ,55 mV and a spike threshold (, ,45 mV). On the other hand, no subthreshold calcium conductances played a significant role in the intrinsic bursting of ventral mossy cells. These observations demonstrate the heterogeneous electrophysiological properties of hilar mossy cells, and suggest that the subthreshold INaP plays a major role in the intrinsic rhythmic bursting of ventral mossy cells. [source] | |||||||||||||||||||||||||||||||