ABSTRACTThe diverse electrical, chemical and structural properties of the functional derivatives ... more ABSTRACTThe diverse electrical, chemical and structural properties of the functional derivatives of carbon nanotubes (CNTs) have shown biomedical possibilities for neuroprosthesis or neural interfaces. However, the studies have been generally confined to metallic CNTs that affect cell viability unless chemically functionalized for biocompatibility. Here, we explored the effects of semiconducting single-walled carbon nanotubes (ssw-CNT), on the active electrical properties of dissociated hippocampal neuronsin-vitrousing multielectrode array, calcium imaging and whole-cell patch clamp recordings. The findings show that ssw-CNT treatment regulates neural network excitability from burst to tonic firing by changing the calcium dynamics. However, at a single neuronal level, ssw-CNT increases neuronal excitability.
Normal anterior pituitary cells, in their diversity and heterogeneity, provide a rich source of m... more Normal anterior pituitary cells, in their diversity and heterogeneity, provide a rich source of models for secretory function. However, until recently they have largely been neglected in favour of neoplastic, clonal tumour cell lines of pituitary origin, which have enabled a number of studies on supposedly homogeneous cell types. Because many of these lines appear to lack key peptide and neurotransmitter receptors, as well as being degranulated with accompanying abnormal levels of secretion, we have developed a range of normal primary anterior pituitary cell cultures using dispersion and enrichment techniques. By studying lactotrophs, somatotrophs and gonadotrophs we have revealed a number of possible transduction mechanisms by which receptors for hypothalamic peptides and neurotransmitters may control secretion. In particular, the transduction events controlling secretion from pituitary cells may differ fundamentally from those found in other cell types. Patch-clamp recordings in these various pituitary cell preparations have revealed substantial populations of voltage-dependent Na + , Ca 2+ and K + channels which may support action potentials in these cells. Although activation of these channels may gate Ca 2+ entry to the cells under some conditions, our evidence taken with that of other laboratories suggests that peptide-receptor interactions leading to hormone secretion occur independently of significant membrane depolarization. Rather, secretion of hormone and rises in intracellular calcium measured with new probes for intracellular calcium activity, can occur in response to hypothalamic peptide activation in the absence of substantial changes in membrane potential. These changes in intracellular calcium activity almost certainly depend on both intracellular and extracellular calcium sources. In addition, strong evidence of a role for multiple intracellular receptors and modulators in the secretory event suggests we should consider the plasma
The whole-cell patch-clamp technique was used to measure cell membrane capacitance (Cm) to monito... more The whole-cell patch-clamp technique was used to measure cell membrane capacitance (Cm) to monitor exocytosis in single-cultured bovine prolactin-secreting cells (lactotrophs) of the anterior pituitary. The cells were dialyzed with solutions containing different concentrations of ionised Ca and non-hydrolyzable GTP analogues (GTP-v-S and GMP-PNP) to activate G-proteins. We have identified two distinct effects of G-protein activation on Ca-induced exocytosis: (i) the maximum Cm increase due to intracellular Ca-dependent exocytosis was diminished, suggesting an inhibitory role of G-proteins close to the site of granule fusion, while (ii) the rate of Cm increase (d Cm/At) was facilitated, revealing conversely a stimulatory role of G-proteins in the translocation of secretory granules to the fusion sites.
Hippocampal neurons are affected by chronic stress and have a high density of cytoplasmic mineral... more Hippocampal neurons are affected by chronic stress and have a high density of cytoplasmic mineralocorticoid and glucocorticoid receptors (MR and GR). Detailed studies on the genomic effects of the stress hormone corticosterone at physiologically relevant concentrations on different steps in synaptic transmission are limited. In this study, we tried to delineate how activation of MR and GR by different concentrations of corticosterone affects synaptic transmission at various levels. The effect of 3-h corticosterone (25, 50, and 100 nM) treatment on depolarization-mediated calcium influx, vesicular release and properties of miniature excitatory post-synaptic currents (mEPSCs) were studied in cultured hippocampal neurons. Activation of MR with 25 nM corticosterone treatment resulted in enhanced depolarization-mediated calcium influx via a transcription-dependent process and increased frequency of mEPSCs with larger amplitude. On the other hand, activation of GR upon 100 nM corticosterone treatment resulted in increase in the rate of vesicular release via the genomic actions of GR. Furthermore, GR activation led to significant increase in the frequency of mEPSCs with larger amplitude and faster decay. Our studies indicate that differential activation of the dual receptor system of MR and GR by corticosterone targets the steps in synaptic transmission differently.
The local fast-spiking interneurons (FSINs) are considered to be crucial for the generation, main... more The local fast-spiking interneurons (FSINs) are considered to be crucial for the generation, maintenance, and modulation of neuronal network oscillations especially in the gamma frequency band. Gamma frequency oscillations have been associated with different aspects of behavior. But the prolonged effects of gamma frequency synaptic activity on the FSINs remain elusive. Using whole cell current clamp patch recordings, we observed a sustained decrease of intrinsic excitability in the FSINs of the dentate gyrus (DG) following repetitive stimulations of the mossy fibers at 30 Hz (gamma bursts). Surprisingly, the granule cells (GCs) did not express intrinsic plastic changes upon similar synaptic excitation of their apical dendritic inputs. Interestingly, pairing the gamma bursts with membrane hyperpolarization accentuated the plasticity in FSINs following the induction protocol, while the plasticity attenuated following gamma bursts paired with membrane depolarization. Paired pulse ratio measurement of the synaptic responses did not show significant changes during the experiments. However, the induction protocols were accompanied with postsynaptic calcium rise in FSINs. Interestingly, the maximum and the minimum increase occurred during gamma bursts with membrane hyperpolarization and depolarization respectively. Including a selective blocker of calcium-permeable AMPA receptors (CP-AMPARs) in the bath; significantly attenuated the calcium rise and blocked the membrane potential dependence of the calcium rise in the FSINs, suggesting their involvement in the observed phenomenon. Chelation of intracellular calcium, blocking HCN channel conductance or blocking CP-AMPARs during the experiment forbade the long lasting expression of the plasticity. Simultaneous dual patch recordings from FSINs and synaptically connected putative GCs confirmed the decreased inhibition in the GCs accompanying the decreased intrinsic excitability in the FSINs. Experimentally constrained network simulations using NEURON predicted increased spiking in the GC owing to decreased input resistance in the FSIN. We hypothesize that the selective plasticity in the FSINs induced by local network activity may serve to increase information throughput into the downstream hippocampal subfields besides providing neuroprotection to the FSINs.
ObjectiveMuch evidence suggests that the subiculum plays a significant role in the regulation of ... more ObjectiveMuch evidence suggests that the subiculum plays a significant role in the regulation of epileptic activity. Lactate acts as a neuroprotective agent against many conditions that cause brain damage. During epileptic seizures, lactate formation reaches up to ~6 mmol/L in the brain. We investigated the effect of lactate on subicular pyramidal neurons after induction of epileptiform activity using 4‐aminopyridine (4‐AP‐0Mg2+) in an in vitro epilepsy model in rats. The signaling mechanism associated with the suppression of epileptiform discharges by lactate was also investigated.MethodsWe used patch clamp electrophysiology recordings on rat subicular neurons of acute hippocampal slices. Immunohistochemistry was used for demonstrating the expression of hydroxycarboxylic acid receptor 1 (HCA1) in the subiculum.ResultsOur study showed that application of 6 mmol/L lactate after induction of epileptiform activity reduced spike frequency (control 2.5 ± 1.23 Hz vs lactate 1.01 ± 0.91 Hz, P = .049) and hyperpolarized the subicular neurons (control −51.8 ± 1.9 mV vs lactate −57.2 ± 3.56 mV, P = .002) in whole cell patch‐clamp experiments. After confirming the expression of HCA1 in subicular neurons, we demonstrated that lactate‐mediated effect occurs via HCA1 by using its specific agonist. All values are mean ±SD. Electrophysiological recordings revealed the involvement of Gβγ and intracellular cAMP in the lactate‐induced effect. Furthermore, current‐clamp and voltage‐clamp experiments showed that the G protein–coupled inwardly rectifying potassium (GIRK) channel blocker tertiapin‐Q, negated the lactate‐induced inhibitory effect, which confirmed that lactate application results in outward GIRK current.SignificanceOur finding points toward the potential role of lactate as an anticonvulsant by showing lactate‐induced suppression of epileptiform activity in subicular neurons. The study gives a different insight by suggesting importance of endogenous metabolite and associated signaling factors, which can aid in improving the present therapeutic approach for treating epilepsy.
Synfire waves are propagating spike packets in synfire chains, which are feedforward chains embed... more Synfire waves are propagating spike packets in synfire chains, which are feedforward chains embedded in random networks. Although synfire waves have proved to be effective quantification for network activity with clear relations to network structure, their utilities are largely limited to feedforward networks with low background activity. To overcome these shortcomings, we describe a novel generalisation of synfire waves, and define 'synconset wave' as a cascade of first spikes within a synchronisation event. Synconset waves would occur in 'synconset chains', which are feedforward chains embedded in possibly heavily recurrent networks with heavy background activity. We probed the utility of synconset waves using simulation of single compartment neuron network models with biophysically realistic conductances, and demonstrated that the spread of synconset waves directly follows from the network connectivity matrix and is modulated by top-down inputs and the resultant oscillations. Such synconset profiles lend intuitive insights into network organisation in terms of connection probabilities between various network regions rather than an adjacency matrix. To test this intuition, we develop a Bayesian likelihood function that quantifies the probability that an observed synfire wave was caused by a given network. Further, we demonstrate it's utility in the inverse problem of identifying the network that caused a given synfire wave. This method was effective even in highly subsampled networks where only a small subset of neurons were accessible, thus showing it's utility in experimental estimation of connectomes in real neuronal-networks. Together, we propose synconset chains/waves as an effective framework for understanding the impact of network structure on function, and as a step towards developing physiology-driven network identification methods. Finally, as synconset chains extend the utilities of synfire chains to arbitrary networks, we suggest utilities of our framework to several aspects of network physiology including cell assemblies, population codes, and oscillatory synchrony.
Pannexins are single-membrane large-pore ion channels that release ATP upon activation. Three iso... more Pannexins are single-membrane large-pore ion channels that release ATP upon activation. Three isoforms of pannexins, 1, 2, and 3, perform diverse cellular roles, including inflammation, differentiation, neuropathic pain, and ATP release. In this study, we report the cryoEM structure of pannexin 3 at 3.9 Å and characterize the structural differences with pannexin isoforms 1 and 2. We observe the organization of the Pannexin3 vestibule into two distinct chambers with a wider pore radius in comparison to both PANX1 and 2 isoforms. We further report the structure of pannexin1 congenital mutant R217H in the resolution range of 3.9 Å. The congenital mutant R217H in transmembrane helix3 (TM3), R217H induce structural changes that leads to a partially closed pore and altered ATP interaction propensities. The channel conductance of the congenital mutant displays weakened voltage sensitivity. The results showcase a complete comparison of the three pannexin isoform structures that along with t...
Besides having a role in signal transduction some trimeric G-proteins may be involved in a late s... more Besides having a role in signal transduction some trimeric G-proteins may be involved in a late stage of exocytosis. Using immunocytochemistry and confocal microscopy we found that Gi3-protein resides mainly in the plasma membrane, whereas Gi1/2-protein is preferentially associated with secretory granules. To study the function of trimeric Gi3- and Gi1/2-proteins, secretory responses in single rat melanotrophs were monitored by patch-clamp membrane capacitance measurements. We report here that mastoparan, an activator of trimeric G-proteins, enhances calcium-induced secretory activity in rat melanotrophs. The introduction of synthetic peptides corresponding to the C-terminal domain of the α-subunit of Gi3- and Gi1/2-proteins indicated that Gi3 peptide specifically blocked the mastoparan-stimulated secretory activity, which indicates an involvement of a trimeric Gi3-protein in mastoparan-stimulated secretory activity. Flash photolysis of caged Ca2+-elicited biphasic capacitance incre...
Fast spiking interneurons (FSINs) have an important role in neuronal network dynamics. Although p... more Fast spiking interneurons (FSINs) have an important role in neuronal network dynamics. Although plasticity of synaptic properties is known to affect network synchrony, the role of plasticity of FSINs' intrinsic excitability on network dynamics remain elusive. Using computational approaches in an excitatory-FSIN model network (EI) based on previously established hippocampal neuronal models we show that altered FSIN intrinsic excitability robustly affects the coherence and frequency of network firing monotonically in the connected excitatory network. Surprisingly, the effect of FSIN excitability was dependent on the mechanisms associated with changes in intrinsic excitability rather than on the direction of the change. Decreasing FSIN excitability by decreasing the membrane specific resistance (R m), increasing peak HCN conductance (g ih bar) increased the excitatory network coherence while increased peak delayed potassium conductance (g KD bar) decreased the coherence. However, the perturbations affected the excitatory network frequency in a similar manner. Further, in an isolated FSIN all-to-all network (II), decreasing FSIN excitability caused significant decrease in the network steady-state frequency due to any of the alterations. However, II network coherence remained unaltered with change in FSIN R m but increased with higher g ih bar and lower g KD bar. Interestingly, decreased FSIN R in could partially rescue the decreasing EI network coherence with increasing g KD bar. The phenomenon of FSIN R m , g ih bar and g KD bar dependent EI network coherence alterations was robust for different proportions of plastic FSINs. Our results indicate that plasticity of intrinsic excitability in FSINs can regulate network dynamics and thus serve as an important network strategy during different physiological states.
We report the observation of a circular differential two-photon photoluminescence (TPPL) response... more We report the observation of a circular differential two-photon photoluminescence (TPPL) response from a threedimensional chiral metamaterial, comprising a system of achiral (spherical) metal nanoparticles arranged on a chiral (helical) dielectric template. The enhanced dipolar response of the individual particles arising from their strong electromagnetic coupling resulted in strong photoluminescence under peak illumination intensities as low as 2 × 10 3 W/cm 2. The TPPL signal was of approximately equal magnitude but of opposite sign, which depended on both the circular polarization state of the incident beam and the handedness of the helical geometry. The strong chiro-optical effect observed in these experiments may be relevant to technologies related to nonlinear plasmonics, in particular imaging applications where control over the polarization state of the imaged photons may be desirable.
We have utilised dynamic video hnaghtg (Magical, Joyce Locbl) on Fura2-loaded pnuactin and growth... more We have utilised dynamic video hnaghtg (Magical, Joyce Locbl) on Fura2-loaded pnuactin and growth hormone-secreting cells to investigate intracelhrlar ionised calchun regulation by peptldes and transmitters acting on intracell~messenger pathways. A flwtescent antibody probe has also been employed to reveal recently released hormone. To investigate exocytosis directly, dynamic measurements of membrane capacitance have been made. These new technologies have demonstmted that prolactin and growth hormone secretion am calcium-dependent, this calcium deriving from both intracellular and extra&Mar sources according to the signal pathway stimulated. CAMP pathways, for instance, appear to regulate intlux across the plasma membrane, whereas IF'3 governs intracellular homeostasis. These signal pathways also interact hi a complementary fashion. Voltage-dependent and independent pathways are important Direct measurements of membrane surface area have revealed that CAMP and non-hydrolysable GTP analogues also modulate Cadependent exocytosis. The site of action is most probably close to the membrane fusion site, and may involve modulation of cytoslceletal integrity as well as membrane fusion. CAMP shifts the 'equilibrium' of membrane turnover towards exocytosis, whereas GTP-r-S, for instance, promotes membrane recovery or inhibitionof exocytosis, both acting on short time scales. Imaging of a fluorescent probe for recently released hormone has suggested these cells may possess specialised plasma membrane sites at which hormone secretion can occur. These release sites may also be correlated with local&d rises in calcium within the cell cytosol.
2015 28th International Conference on VLSI Design, 2015
Neuro-electronic hybrid systems have been gaining interest of researchers as a possible architect... more Neuro-electronic hybrid systems have been gaining interest of researchers as a possible architecture for computing. This aims to exploit the strengths of biological neuronal systems with their immense parallel processing and learning capabilities along with that of VLSI systems. Towards this end, we have set up a system which demonstrates the use of a live neuronal culture to solve a real world problem of controlling a robot doing the task of obstacle avoidance. We show that a neuronal culture can look at the sensor inputs to the robot and generate motor commands to allow it to explore an arena while avoiding obstacles.
Proceedings of the National Academy of Sciences, 2000
Although many proteins essential for regulated neurotransmitter and peptide hormone secretion hav... more Although many proteins essential for regulated neurotransmitter and peptide hormone secretion have been identified, little is understood about their precise roles at specific stages of the multistep pathway of exocytosis. To study the function of CAPS (Ca 2+ -dependent activator protein for secretion), a protein required for Ca 2+ -dependent exocytosis of dense-core vesicles, secretory responses in single rat melanotrophs were monitored by patch-clamp membrane capacitance measurements. Flash photolysis of caged Ca 2+ elicited biphasic capacitance increases consisting of rapid and slow components with distinct Ca 2+ dependencies. A threshold of ≈10 μM Ca 2+ was required to trigger the slow component, while the rapid capacitance increase was recorded already at a intracellular Ca 2+ activity < 10 μM. Both kinetic membrane capacitance components were abolished by botulinum neurotoxin B or E treatment, suggesting involvement of SNARE (soluble N -ethylmaleimide-sensitive factor attach...
L-PGlu-(2-propyl)-L-His-L-ProNH 2 (NP-647) is a CNS active thyrotropin-releasing hormone (TRH) an... more L-PGlu-(2-propyl)-L-His-L-ProNH 2 (NP-647) is a CNS active thyrotropin-releasing hormone (TRH) analog with potential application in various CNS disorders including seizures. In the present study, mechanism of action for protective effect of NP-647 was explored by studying role of NP-647 on epileptiform activity and sodium channels by using patchclamp methods. Epileptiform activity was induced in subicular pyramidal neurons of hippocampal slice of rat by perfusing 4-aminopyridine (4-AP) containing Mg ؉2-free normal artificial cerebrospinal fluid (nACSF). Increase in mean firing frequency was observed after perfusion of 4-AP and zero Mg ؉2 (2.10؎0.47 Hz) as compared with nACSF (0.12؎0.08 Hz). A significant decrease in mean firing frequency (0.61؎0.22 Hz), mean frequency of epileptiform events (0.03؎0.02 Hz vs. 0.22؎0.05 Hz of 4-AP؉0 Mg), and average number of action potentials in paroxysmal depolarization shift-burst (2.54؎1.21 Hz vs. 8.16؎0.88 Hz of 4-AP؉0 Mg) was observed. A significant reduction in peak dV/dt (246؎19 mV ms ؊1 vs. 297؎18 mV ms ؊1 of 4-AP؉0 Mg) and increase (1.332؎0.018 ms vs. 1.292؎0.019 ms of 4-AP؉0 Mg) in time required to reach maximum depolarization were observed indicating role of sodium channels. Concentration-dependent depression of sodium current was observed after exposure to dorsal root ganglion neurons to NP-647. NP-647 at different concentrations (1, 3, and 10 M) depressed sodium current (15؎0.5%, 50؎2.6%, and 75؎0.7%, respectively). However, NP-647 did not show change in the peak sodium current in CNa18 cells. Results of present study demonstrated potential of NP-647 in the inhibition of epileptiform activity by inhibiting sodium channels indirectly.
The three-dimensional (3D) NMR solution structure (MeOH) of the highly hydrophobic delta-conotoxi... more The three-dimensional (3D) NMR solution structure (MeOH) of the highly hydrophobic delta-conotoxin delta-Am2766 from the molluscivorous snail Conus amadis has been determined. Fifteen converged structures were obtained on the basis of 262 distance constraints, 25 torsion-angle constraints, and ten constraints based on disulfide linkages and H-bonds. The root-mean-square deviations (rmsd) about the averaged coordinates of the backbone (N, C(alpha), C) and (all) heavy atoms were 0.62+/-0.20 and 1.12+/-0.23 A, respectively. The structures determined are of good stereochemical quality, as evidenced by the high percentage (100%) of backbone dihedral angles that occupy favorable and additionally allowed regions of the Ramachandran map. The structure of delta-Am2766 consists of a triple-stranded antiparallel beta-sheet, and of four turns. The three disulfides form the classical &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;inhibitory cysteine knot&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39; motif. So far, only one tertiary structure of a delta-conotoxin has been reported; thus, the tertiary structure of delta-Am2766 is the second such example. Another Conus peptide, Am2735 from C. amadis, has also been purified and sequenced. Am2735 shares 96% sequence identity with delta-Am2766. Unlike delta-Am2766, Am2735 does not inhibit the fast inactivation of Na+ currents in rat brain Na(v)1.2 Na+ channels at concentrations up to 200 nM.
The interaction of zwitterionic lipid DMPC and DPPC with cyclic hexapeptide, cyclo (D-Ala-L-Pro-L... more The interaction of zwitterionic lipid DMPC and DPPC with cyclic hexapeptide, cyclo (D-Ala-L-Pro-L-Ala)2 was studied using circular dichroism (CD) and differential scanning calorimetry (DSC). Preliminary membrane conductance results showed that the peptide has a tendency to form channels inside the lipid bilayer. CD studies indicated that as the lipid/peptide (L/P) ratio (DMPC/peptide) was increased, the magnitude of the negative CD band having a lambda(max) around 200 nm decreased. At a L/P ratio of 210:1, this band disappeared completely, indicating dramatic conformational changes in the peptide on interaction with the lipid bilayer. Reduction of the phase transition temperature and the maximum heat capacity of the lipid bilayer (DPPC) for gel-to-liquid crystalline phase transition indicates a strong interaction of the peptide with the lipid bilayer.
Tetrapentylammonium (TPeA) block of rat brain type IIA sodium channel α subunit was studied using... more Tetrapentylammonium (TPeA) block of rat brain type IIA sodium channel α subunit was studied using whole cell patch clamp. Results indicate that TPeA blocks the inactivating brain sodium channel in a potential and use‐dependent manner similar to that of the cardiac sodium channel. Removal of inactivation using chloramine‐T (CT) unmasks a time‐dependent block by TPeA consistent with slow blocking kinetics. On the other hand, no time dependence is observed when inactivation is abolished by modification with veratridine. TPeA does not bind in a potential‐dependent fashion to veratridine‐modified channels and does not significantly affect gating of veratridine‐modified channels suggesting that high affinity binding of TPeA to the brain sodium channel is lost after veratridine modification.British Journal of Pharmacology (2001) 132, 1755–1760; doi:10.1038/sj.bjp.0703973
ABSTRACTThe diverse electrical, chemical and structural properties of the functional derivatives ... more ABSTRACTThe diverse electrical, chemical and structural properties of the functional derivatives of carbon nanotubes (CNTs) have shown biomedical possibilities for neuroprosthesis or neural interfaces. However, the studies have been generally confined to metallic CNTs that affect cell viability unless chemically functionalized for biocompatibility. Here, we explored the effects of semiconducting single-walled carbon nanotubes (ssw-CNT), on the active electrical properties of dissociated hippocampal neuronsin-vitrousing multielectrode array, calcium imaging and whole-cell patch clamp recordings. The findings show that ssw-CNT treatment regulates neural network excitability from burst to tonic firing by changing the calcium dynamics. However, at a single neuronal level, ssw-CNT increases neuronal excitability.
Normal anterior pituitary cells, in their diversity and heterogeneity, provide a rich source of m... more Normal anterior pituitary cells, in their diversity and heterogeneity, provide a rich source of models for secretory function. However, until recently they have largely been neglected in favour of neoplastic, clonal tumour cell lines of pituitary origin, which have enabled a number of studies on supposedly homogeneous cell types. Because many of these lines appear to lack key peptide and neurotransmitter receptors, as well as being degranulated with accompanying abnormal levels of secretion, we have developed a range of normal primary anterior pituitary cell cultures using dispersion and enrichment techniques. By studying lactotrophs, somatotrophs and gonadotrophs we have revealed a number of possible transduction mechanisms by which receptors for hypothalamic peptides and neurotransmitters may control secretion. In particular, the transduction events controlling secretion from pituitary cells may differ fundamentally from those found in other cell types. Patch-clamp recordings in these various pituitary cell preparations have revealed substantial populations of voltage-dependent Na + , Ca 2+ and K + channels which may support action potentials in these cells. Although activation of these channels may gate Ca 2+ entry to the cells under some conditions, our evidence taken with that of other laboratories suggests that peptide-receptor interactions leading to hormone secretion occur independently of significant membrane depolarization. Rather, secretion of hormone and rises in intracellular calcium measured with new probes for intracellular calcium activity, can occur in response to hypothalamic peptide activation in the absence of substantial changes in membrane potential. These changes in intracellular calcium activity almost certainly depend on both intracellular and extracellular calcium sources. In addition, strong evidence of a role for multiple intracellular receptors and modulators in the secretory event suggests we should consider the plasma
The whole-cell patch-clamp technique was used to measure cell membrane capacitance (Cm) to monito... more The whole-cell patch-clamp technique was used to measure cell membrane capacitance (Cm) to monitor exocytosis in single-cultured bovine prolactin-secreting cells (lactotrophs) of the anterior pituitary. The cells were dialyzed with solutions containing different concentrations of ionised Ca and non-hydrolyzable GTP analogues (GTP-v-S and GMP-PNP) to activate G-proteins. We have identified two distinct effects of G-protein activation on Ca-induced exocytosis: (i) the maximum Cm increase due to intracellular Ca-dependent exocytosis was diminished, suggesting an inhibitory role of G-proteins close to the site of granule fusion, while (ii) the rate of Cm increase (d Cm/At) was facilitated, revealing conversely a stimulatory role of G-proteins in the translocation of secretory granules to the fusion sites.
Hippocampal neurons are affected by chronic stress and have a high density of cytoplasmic mineral... more Hippocampal neurons are affected by chronic stress and have a high density of cytoplasmic mineralocorticoid and glucocorticoid receptors (MR and GR). Detailed studies on the genomic effects of the stress hormone corticosterone at physiologically relevant concentrations on different steps in synaptic transmission are limited. In this study, we tried to delineate how activation of MR and GR by different concentrations of corticosterone affects synaptic transmission at various levels. The effect of 3-h corticosterone (25, 50, and 100 nM) treatment on depolarization-mediated calcium influx, vesicular release and properties of miniature excitatory post-synaptic currents (mEPSCs) were studied in cultured hippocampal neurons. Activation of MR with 25 nM corticosterone treatment resulted in enhanced depolarization-mediated calcium influx via a transcription-dependent process and increased frequency of mEPSCs with larger amplitude. On the other hand, activation of GR upon 100 nM corticosterone treatment resulted in increase in the rate of vesicular release via the genomic actions of GR. Furthermore, GR activation led to significant increase in the frequency of mEPSCs with larger amplitude and faster decay. Our studies indicate that differential activation of the dual receptor system of MR and GR by corticosterone targets the steps in synaptic transmission differently.
The local fast-spiking interneurons (FSINs) are considered to be crucial for the generation, main... more The local fast-spiking interneurons (FSINs) are considered to be crucial for the generation, maintenance, and modulation of neuronal network oscillations especially in the gamma frequency band. Gamma frequency oscillations have been associated with different aspects of behavior. But the prolonged effects of gamma frequency synaptic activity on the FSINs remain elusive. Using whole cell current clamp patch recordings, we observed a sustained decrease of intrinsic excitability in the FSINs of the dentate gyrus (DG) following repetitive stimulations of the mossy fibers at 30 Hz (gamma bursts). Surprisingly, the granule cells (GCs) did not express intrinsic plastic changes upon similar synaptic excitation of their apical dendritic inputs. Interestingly, pairing the gamma bursts with membrane hyperpolarization accentuated the plasticity in FSINs following the induction protocol, while the plasticity attenuated following gamma bursts paired with membrane depolarization. Paired pulse ratio measurement of the synaptic responses did not show significant changes during the experiments. However, the induction protocols were accompanied with postsynaptic calcium rise in FSINs. Interestingly, the maximum and the minimum increase occurred during gamma bursts with membrane hyperpolarization and depolarization respectively. Including a selective blocker of calcium-permeable AMPA receptors (CP-AMPARs) in the bath; significantly attenuated the calcium rise and blocked the membrane potential dependence of the calcium rise in the FSINs, suggesting their involvement in the observed phenomenon. Chelation of intracellular calcium, blocking HCN channel conductance or blocking CP-AMPARs during the experiment forbade the long lasting expression of the plasticity. Simultaneous dual patch recordings from FSINs and synaptically connected putative GCs confirmed the decreased inhibition in the GCs accompanying the decreased intrinsic excitability in the FSINs. Experimentally constrained network simulations using NEURON predicted increased spiking in the GC owing to decreased input resistance in the FSIN. We hypothesize that the selective plasticity in the FSINs induced by local network activity may serve to increase information throughput into the downstream hippocampal subfields besides providing neuroprotection to the FSINs.
ObjectiveMuch evidence suggests that the subiculum plays a significant role in the regulation of ... more ObjectiveMuch evidence suggests that the subiculum plays a significant role in the regulation of epileptic activity. Lactate acts as a neuroprotective agent against many conditions that cause brain damage. During epileptic seizures, lactate formation reaches up to ~6 mmol/L in the brain. We investigated the effect of lactate on subicular pyramidal neurons after induction of epileptiform activity using 4‐aminopyridine (4‐AP‐0Mg2+) in an in vitro epilepsy model in rats. The signaling mechanism associated with the suppression of epileptiform discharges by lactate was also investigated.MethodsWe used patch clamp electrophysiology recordings on rat subicular neurons of acute hippocampal slices. Immunohistochemistry was used for demonstrating the expression of hydroxycarboxylic acid receptor 1 (HCA1) in the subiculum.ResultsOur study showed that application of 6 mmol/L lactate after induction of epileptiform activity reduced spike frequency (control 2.5 ± 1.23 Hz vs lactate 1.01 ± 0.91 Hz, P = .049) and hyperpolarized the subicular neurons (control −51.8 ± 1.9 mV vs lactate −57.2 ± 3.56 mV, P = .002) in whole cell patch‐clamp experiments. After confirming the expression of HCA1 in subicular neurons, we demonstrated that lactate‐mediated effect occurs via HCA1 by using its specific agonist. All values are mean ±SD. Electrophysiological recordings revealed the involvement of Gβγ and intracellular cAMP in the lactate‐induced effect. Furthermore, current‐clamp and voltage‐clamp experiments showed that the G protein–coupled inwardly rectifying potassium (GIRK) channel blocker tertiapin‐Q, negated the lactate‐induced inhibitory effect, which confirmed that lactate application results in outward GIRK current.SignificanceOur finding points toward the potential role of lactate as an anticonvulsant by showing lactate‐induced suppression of epileptiform activity in subicular neurons. The study gives a different insight by suggesting importance of endogenous metabolite and associated signaling factors, which can aid in improving the present therapeutic approach for treating epilepsy.
Synfire waves are propagating spike packets in synfire chains, which are feedforward chains embed... more Synfire waves are propagating spike packets in synfire chains, which are feedforward chains embedded in random networks. Although synfire waves have proved to be effective quantification for network activity with clear relations to network structure, their utilities are largely limited to feedforward networks with low background activity. To overcome these shortcomings, we describe a novel generalisation of synfire waves, and define 'synconset wave' as a cascade of first spikes within a synchronisation event. Synconset waves would occur in 'synconset chains', which are feedforward chains embedded in possibly heavily recurrent networks with heavy background activity. We probed the utility of synconset waves using simulation of single compartment neuron network models with biophysically realistic conductances, and demonstrated that the spread of synconset waves directly follows from the network connectivity matrix and is modulated by top-down inputs and the resultant oscillations. Such synconset profiles lend intuitive insights into network organisation in terms of connection probabilities between various network regions rather than an adjacency matrix. To test this intuition, we develop a Bayesian likelihood function that quantifies the probability that an observed synfire wave was caused by a given network. Further, we demonstrate it's utility in the inverse problem of identifying the network that caused a given synfire wave. This method was effective even in highly subsampled networks where only a small subset of neurons were accessible, thus showing it's utility in experimental estimation of connectomes in real neuronal-networks. Together, we propose synconset chains/waves as an effective framework for understanding the impact of network structure on function, and as a step towards developing physiology-driven network identification methods. Finally, as synconset chains extend the utilities of synfire chains to arbitrary networks, we suggest utilities of our framework to several aspects of network physiology including cell assemblies, population codes, and oscillatory synchrony.
Pannexins are single-membrane large-pore ion channels that release ATP upon activation. Three iso... more Pannexins are single-membrane large-pore ion channels that release ATP upon activation. Three isoforms of pannexins, 1, 2, and 3, perform diverse cellular roles, including inflammation, differentiation, neuropathic pain, and ATP release. In this study, we report the cryoEM structure of pannexin 3 at 3.9 Å and characterize the structural differences with pannexin isoforms 1 and 2. We observe the organization of the Pannexin3 vestibule into two distinct chambers with a wider pore radius in comparison to both PANX1 and 2 isoforms. We further report the structure of pannexin1 congenital mutant R217H in the resolution range of 3.9 Å. The congenital mutant R217H in transmembrane helix3 (TM3), R217H induce structural changes that leads to a partially closed pore and altered ATP interaction propensities. The channel conductance of the congenital mutant displays weakened voltage sensitivity. The results showcase a complete comparison of the three pannexin isoform structures that along with t...
Besides having a role in signal transduction some trimeric G-proteins may be involved in a late s... more Besides having a role in signal transduction some trimeric G-proteins may be involved in a late stage of exocytosis. Using immunocytochemistry and confocal microscopy we found that Gi3-protein resides mainly in the plasma membrane, whereas Gi1/2-protein is preferentially associated with secretory granules. To study the function of trimeric Gi3- and Gi1/2-proteins, secretory responses in single rat melanotrophs were monitored by patch-clamp membrane capacitance measurements. We report here that mastoparan, an activator of trimeric G-proteins, enhances calcium-induced secretory activity in rat melanotrophs. The introduction of synthetic peptides corresponding to the C-terminal domain of the α-subunit of Gi3- and Gi1/2-proteins indicated that Gi3 peptide specifically blocked the mastoparan-stimulated secretory activity, which indicates an involvement of a trimeric Gi3-protein in mastoparan-stimulated secretory activity. Flash photolysis of caged Ca2+-elicited biphasic capacitance incre...
Fast spiking interneurons (FSINs) have an important role in neuronal network dynamics. Although p... more Fast spiking interneurons (FSINs) have an important role in neuronal network dynamics. Although plasticity of synaptic properties is known to affect network synchrony, the role of plasticity of FSINs' intrinsic excitability on network dynamics remain elusive. Using computational approaches in an excitatory-FSIN model network (EI) based on previously established hippocampal neuronal models we show that altered FSIN intrinsic excitability robustly affects the coherence and frequency of network firing monotonically in the connected excitatory network. Surprisingly, the effect of FSIN excitability was dependent on the mechanisms associated with changes in intrinsic excitability rather than on the direction of the change. Decreasing FSIN excitability by decreasing the membrane specific resistance (R m), increasing peak HCN conductance (g ih bar) increased the excitatory network coherence while increased peak delayed potassium conductance (g KD bar) decreased the coherence. However, the perturbations affected the excitatory network frequency in a similar manner. Further, in an isolated FSIN all-to-all network (II), decreasing FSIN excitability caused significant decrease in the network steady-state frequency due to any of the alterations. However, II network coherence remained unaltered with change in FSIN R m but increased with higher g ih bar and lower g KD bar. Interestingly, decreased FSIN R in could partially rescue the decreasing EI network coherence with increasing g KD bar. The phenomenon of FSIN R m , g ih bar and g KD bar dependent EI network coherence alterations was robust for different proportions of plastic FSINs. Our results indicate that plasticity of intrinsic excitability in FSINs can regulate network dynamics and thus serve as an important network strategy during different physiological states.
We report the observation of a circular differential two-photon photoluminescence (TPPL) response... more We report the observation of a circular differential two-photon photoluminescence (TPPL) response from a threedimensional chiral metamaterial, comprising a system of achiral (spherical) metal nanoparticles arranged on a chiral (helical) dielectric template. The enhanced dipolar response of the individual particles arising from their strong electromagnetic coupling resulted in strong photoluminescence under peak illumination intensities as low as 2 × 10 3 W/cm 2. The TPPL signal was of approximately equal magnitude but of opposite sign, which depended on both the circular polarization state of the incident beam and the handedness of the helical geometry. The strong chiro-optical effect observed in these experiments may be relevant to technologies related to nonlinear plasmonics, in particular imaging applications where control over the polarization state of the imaged photons may be desirable.
We have utilised dynamic video hnaghtg (Magical, Joyce Locbl) on Fura2-loaded pnuactin and growth... more We have utilised dynamic video hnaghtg (Magical, Joyce Locbl) on Fura2-loaded pnuactin and growth hormone-secreting cells to investigate intracelhrlar ionised calchun regulation by peptldes and transmitters acting on intracell~messenger pathways. A flwtescent antibody probe has also been employed to reveal recently released hormone. To investigate exocytosis directly, dynamic measurements of membrane capacitance have been made. These new technologies have demonstmted that prolactin and growth hormone secretion am calcium-dependent, this calcium deriving from both intracellular and extra&Mar sources according to the signal pathway stimulated. CAMP pathways, for instance, appear to regulate intlux across the plasma membrane, whereas IF'3 governs intracellular homeostasis. These signal pathways also interact hi a complementary fashion. Voltage-dependent and independent pathways are important Direct measurements of membrane surface area have revealed that CAMP and non-hydrolysable GTP analogues also modulate Cadependent exocytosis. The site of action is most probably close to the membrane fusion site, and may involve modulation of cytoslceletal integrity as well as membrane fusion. CAMP shifts the 'equilibrium' of membrane turnover towards exocytosis, whereas GTP-r-S, for instance, promotes membrane recovery or inhibitionof exocytosis, both acting on short time scales. Imaging of a fluorescent probe for recently released hormone has suggested these cells may possess specialised plasma membrane sites at which hormone secretion can occur. These release sites may also be correlated with local&d rises in calcium within the cell cytosol.
2015 28th International Conference on VLSI Design, 2015
Neuro-electronic hybrid systems have been gaining interest of researchers as a possible architect... more Neuro-electronic hybrid systems have been gaining interest of researchers as a possible architecture for computing. This aims to exploit the strengths of biological neuronal systems with their immense parallel processing and learning capabilities along with that of VLSI systems. Towards this end, we have set up a system which demonstrates the use of a live neuronal culture to solve a real world problem of controlling a robot doing the task of obstacle avoidance. We show that a neuronal culture can look at the sensor inputs to the robot and generate motor commands to allow it to explore an arena while avoiding obstacles.
Proceedings of the National Academy of Sciences, 2000
Although many proteins essential for regulated neurotransmitter and peptide hormone secretion hav... more Although many proteins essential for regulated neurotransmitter and peptide hormone secretion have been identified, little is understood about their precise roles at specific stages of the multistep pathway of exocytosis. To study the function of CAPS (Ca 2+ -dependent activator protein for secretion), a protein required for Ca 2+ -dependent exocytosis of dense-core vesicles, secretory responses in single rat melanotrophs were monitored by patch-clamp membrane capacitance measurements. Flash photolysis of caged Ca 2+ elicited biphasic capacitance increases consisting of rapid and slow components with distinct Ca 2+ dependencies. A threshold of ≈10 μM Ca 2+ was required to trigger the slow component, while the rapid capacitance increase was recorded already at a intracellular Ca 2+ activity < 10 μM. Both kinetic membrane capacitance components were abolished by botulinum neurotoxin B or E treatment, suggesting involvement of SNARE (soluble N -ethylmaleimide-sensitive factor attach...
L-PGlu-(2-propyl)-L-His-L-ProNH 2 (NP-647) is a CNS active thyrotropin-releasing hormone (TRH) an... more L-PGlu-(2-propyl)-L-His-L-ProNH 2 (NP-647) is a CNS active thyrotropin-releasing hormone (TRH) analog with potential application in various CNS disorders including seizures. In the present study, mechanism of action for protective effect of NP-647 was explored by studying role of NP-647 on epileptiform activity and sodium channels by using patchclamp methods. Epileptiform activity was induced in subicular pyramidal neurons of hippocampal slice of rat by perfusing 4-aminopyridine (4-AP) containing Mg ؉2-free normal artificial cerebrospinal fluid (nACSF). Increase in mean firing frequency was observed after perfusion of 4-AP and zero Mg ؉2 (2.10؎0.47 Hz) as compared with nACSF (0.12؎0.08 Hz). A significant decrease in mean firing frequency (0.61؎0.22 Hz), mean frequency of epileptiform events (0.03؎0.02 Hz vs. 0.22؎0.05 Hz of 4-AP؉0 Mg), and average number of action potentials in paroxysmal depolarization shift-burst (2.54؎1.21 Hz vs. 8.16؎0.88 Hz of 4-AP؉0 Mg) was observed. A significant reduction in peak dV/dt (246؎19 mV ms ؊1 vs. 297؎18 mV ms ؊1 of 4-AP؉0 Mg) and increase (1.332؎0.018 ms vs. 1.292؎0.019 ms of 4-AP؉0 Mg) in time required to reach maximum depolarization were observed indicating role of sodium channels. Concentration-dependent depression of sodium current was observed after exposure to dorsal root ganglion neurons to NP-647. NP-647 at different concentrations (1, 3, and 10 M) depressed sodium current (15؎0.5%, 50؎2.6%, and 75؎0.7%, respectively). However, NP-647 did not show change in the peak sodium current in CNa18 cells. Results of present study demonstrated potential of NP-647 in the inhibition of epileptiform activity by inhibiting sodium channels indirectly.
The three-dimensional (3D) NMR solution structure (MeOH) of the highly hydrophobic delta-conotoxi... more The three-dimensional (3D) NMR solution structure (MeOH) of the highly hydrophobic delta-conotoxin delta-Am2766 from the molluscivorous snail Conus amadis has been determined. Fifteen converged structures were obtained on the basis of 262 distance constraints, 25 torsion-angle constraints, and ten constraints based on disulfide linkages and H-bonds. The root-mean-square deviations (rmsd) about the averaged coordinates of the backbone (N, C(alpha), C) and (all) heavy atoms were 0.62+/-0.20 and 1.12+/-0.23 A, respectively. The structures determined are of good stereochemical quality, as evidenced by the high percentage (100%) of backbone dihedral angles that occupy favorable and additionally allowed regions of the Ramachandran map. The structure of delta-Am2766 consists of a triple-stranded antiparallel beta-sheet, and of four turns. The three disulfides form the classical &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;inhibitory cysteine knot&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39; motif. So far, only one tertiary structure of a delta-conotoxin has been reported; thus, the tertiary structure of delta-Am2766 is the second such example. Another Conus peptide, Am2735 from C. amadis, has also been purified and sequenced. Am2735 shares 96% sequence identity with delta-Am2766. Unlike delta-Am2766, Am2735 does not inhibit the fast inactivation of Na+ currents in rat brain Na(v)1.2 Na+ channels at concentrations up to 200 nM.
The interaction of zwitterionic lipid DMPC and DPPC with cyclic hexapeptide, cyclo (D-Ala-L-Pro-L... more The interaction of zwitterionic lipid DMPC and DPPC with cyclic hexapeptide, cyclo (D-Ala-L-Pro-L-Ala)2 was studied using circular dichroism (CD) and differential scanning calorimetry (DSC). Preliminary membrane conductance results showed that the peptide has a tendency to form channels inside the lipid bilayer. CD studies indicated that as the lipid/peptide (L/P) ratio (DMPC/peptide) was increased, the magnitude of the negative CD band having a lambda(max) around 200 nm decreased. At a L/P ratio of 210:1, this band disappeared completely, indicating dramatic conformational changes in the peptide on interaction with the lipid bilayer. Reduction of the phase transition temperature and the maximum heat capacity of the lipid bilayer (DPPC) for gel-to-liquid crystalline phase transition indicates a strong interaction of the peptide with the lipid bilayer.
Tetrapentylammonium (TPeA) block of rat brain type IIA sodium channel α subunit was studied using... more Tetrapentylammonium (TPeA) block of rat brain type IIA sodium channel α subunit was studied using whole cell patch clamp. Results indicate that TPeA blocks the inactivating brain sodium channel in a potential and use‐dependent manner similar to that of the cardiac sodium channel. Removal of inactivation using chloramine‐T (CT) unmasks a time‐dependent block by TPeA consistent with slow blocking kinetics. On the other hand, no time dependence is observed when inactivation is abolished by modification with veratridine. TPeA does not bind in a potential‐dependent fashion to veratridine‐modified channels and does not significantly affect gating of veratridine‐modified channels suggesting that high affinity binding of TPeA to the brain sodium channel is lost after veratridine modification.British Journal of Pharmacology (2001) 132, 1755–1760; doi:10.1038/sj.bjp.0703973
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Papers by Sujit Sikdar