Insights into channel function via channel dysfunction

The nicotinic synapse has been a touchstone for advances in neuroscience ever since Jean Nicot, the French ambassador to Portugal, sent some tobacco seeds home to Paris in 1550 with a note that the New World plant had interesting effects when smoked. Now the muscle nicotinic acetylcholine receptor (nAChR) is a well-studied example of ligand-gated ion channels. After a motor neuron is stimulated, the nerve impulse reaches the presynaptic terminal, where it evokes release of acetylcholine (ACh) into the synapse. The nAChR depolarizes the postsynaptic muscle and triggers muscle action potentials; muscle contraction follows. To date, several nAChR subtypes have been successfully isolated, purified, imaged, and expressed, and unitary currents have been recorded from these channels (1). Researchers continue to unravel the molecular mechanisms of these macromolecules that are embedded in membranes at vertebrate nerve-muscle synapses, at invertebrate nicotinic synapses (which explains why nicotine-producing tobacco plants have a select advantage against invertebrate pests), and in the vertebrate central system (which explains Jean Nicot’s fascination with those leaves). However, the precise structural events that trigger channel opening or "gating" remain mostly unknown

Intrinsic Gating Properties of a Cloned G Protein-activated Inward Rectifier K^+ Channel

The voltage-, time-, and K^+-dependent properties of a G protein-activated inwardly rectifying K^+ channel (GIRK1/KGA/Kir3.1) cloned from rat atrium were studied in Xenopus oocytes under two-electrode voltage clamp. During maintained G protein activation and in the presence of high external K^+ (V_K = 0 mV), voltage jumps from V_K to negative membrane potentials activated inward GIRK1 K^+ currents with three distinct time-resolved current components. GIRK1 current activation consisted of an instantaneous component that was followed by two components with time constants T_f~50 ms and T_s~400 ms. These activation time constants were weakly voltage dependent, increasing approximately twofold with maximal hyperpolarization from V_K. Voltage-dependent GIRK1 availability, revealed by tail currents at -80 mV after long prepulses, was greatest at potentials negative to V_K and declined to a plateau of approximately half the maximal level at positive voltages. Voltage-dependent GIRK1 availability shifted with V_K and was half maximal at V_K -20 mV; the equivalent gating charge was ~1.6 e^-. The voltage-dependent gating parameters of GIRK1 did not significantly differ for G protein activation by three heterologously expressed signaling pathways: m2 muscarinic receptors, serotonin 1A receptors, or G protein β1y2 subunits. Voltage dependence was also unaffected by agonist concentration. These results indicate that the voltage-dependent gating properties of GIRK1 are not due to extrinsic factors such as agonist-receptor interactions and G protein-channel coupling, but instead are analogous to the intrinsic gating behaviors of other inwardly rectifying K^+ channels

Consumption Growth and Agricultural Shocks in Rural Madagascar

The aim of this paper is to evaluate the effect of rainfall and agricultural shocks on consumption growth in Madagascar. We are also interested in the impact of local endowments in infrastructures and social services on consumption growth. To achieve this goal, a micro model of household consumption growth is estimated thanks to household panel data collected by the Reseau des Observatoires Ruraux (ROR) between 1999 and 2004. Additional data sources include the 2001 communes census organized by the Ilo program of Cornell University. Altogether these different data sources make an unusually rich data set, at least when considered with developing country standards. We use panel data fixed effect estimation technique to remove unobserved household and community level time invariant heterogeneity. We find that production shocks have a substantial impact on consumption growth and we find sign of persistence of rainfall shocks. Roads and education seems to improve household’s consumption growth and remotness decreases it.risks, growth, poverty, Food Security and Poverty,

Rates and Equilibria for a Photoisomerizable Antagonist at the Acetylcholine Receptor of Electrophorus Electroplaques

Voltage-jump and light-flash experiments have been performed on isolated Electrophorus electroplaques exposed simultaneously to nicotinic agonists and to the photoisomerizable compound 2,2'-bis-[α-(trimethylammonium)methyl]-azobenzene (2BQ). Dose-response curves are shifted to the right in a nearly parallel fashion by 2BQ, which suggests competitive antagonism; dose-ratio analyses show apparent dissociation constants of 0.3 and 1 µM for the cis and trans isomers, respectively. Flash-induced trans → cis concentration jumps produce the expected decrease in agonist-induced conductance; the time constant is several tens of milliseconds. From the concentration dependence of these rates, we conclude that the association and dissociation rate constants for the cis-2BQ-receptor binding are approximately ~ 10^8 M^(-1) s^(-1) and 60 s^(-1) at 20ºC; the Q_(10) is 3. Flash-induced cis → trans photoisomerizations produce molecular rearrangements of the ligand-receptor complex, but the resulting relaxations probably reflect the kinetics of buffered diffusion rather than of the interaction between trans-2BQ and the receptor. Antagonists seem to bind about an order of magnitude more slowly than agonists at nicotinic receptors

Expression of an atrial G-protein-activated potassium channel in Xenopus oocytes

Injection of rat atrial RNA into Xenopus oocytes resulted in the expression of a guanine nucleotide binding (G) protein-activated K+ channel. Current through the channel could be activated by acetylcholine or, if RNA encoding a neuronal 5HT1A receptor was coinjected with atrial RNA, by serotonin (5HT). A 5HT-evoked current (I5HT) was observed in oocytes injected with ventricle RNA fractions (of 2.5-5.5 kb) and 5HT1A receptor RNA. I5HT displayed strong inward rectification with very little conductance above the K+ equilibrium potential, was highly selective for K+ over Na+, and was blocked by 5-300 µM Ba2+. I5HT was suppressed by intracellular injection of the nonhydrolyzable analog of GDP, guanosine 5'-[ß-thio]diphosphate, but not by treatment with pertussis toxin (PTX), suggesting coupling of the receptor to the G-protein-activated K+ channel via a PTX-insensitive G protein, possibly endogenously present in the oocyte. Coexpression of the subunit of a PTX-sensitive G protein, Gi2, rendered I5HT sensitive to PTX inhibition. Native oocytes displayed a constitutively active inwardly rectifying K+ current with a lower sensitivity to Ba2+ block; expression of a similar current was also directed by atrial or ventricle RNA of 1.5-3 kb. Xenopus oocytes may be employed for cloning of the G-protein-activated K+ channel cDNA and for studying the coupling between this channel and G proteins

A covalently bound photoisomerizable agonist. Comparison with reversibly bound agonists at electrophorus electroplaques

After disulphide bonds are reduced with dithiothreitol, trans-3-(alpha-bromomethyl)-3’-[alpha-(trimethylammonium)methyl]azobenzene (trans-QBr) alkylates a sulfhydryl group on receptors. The membrane conductance induced by this “tethered agonist” shares many properties with that induced by reversible agonists. Equilibrium conductance increases as the membrane potential is made more negative; the voltage sensitivity resembles that seen with 50 [mu]M carbachol. Voltage- jump relaxations follow an exponential time-course; the rate constants are about twice as large as those seen with 50 mu M carbachol and have the same voltage and temperature sensitivity. With reversible agonists, the rate of channel opening increases with the frequency of agonist-receptor collisions: with tethered trans-Qbr, this rate depends only on intramolecular events. In comparison to the conductance induced by reversible agonists, the QBr-induced conductance is at least 10-fold less sensitive to competitive blockade by tubocurarine and roughly as sensitive to “open-channel blockade” bu QX-222. Light-flash experiments with tethered QBr resemble those with the reversible photoisomerizable agonist, 3,3’,bis-[alpha-(trimethylammonium)methyl]azobenzene (Bis-Q): the conductance is increased by cis {arrow} trans photoisomerizations and decreased by trans {arrow} cis photoisomerizations. As with Bis-Q, ligh-flash relaxations have the same rate constant as voltage-jump relaxations. Receptors with tethered trans isomer. By comparing the agonist-induced conductance with the cis/tans ratio, we conclude that each channel’s activation is determined by the configuration of a single tethered QBr molecule. The QBr-induced conductance shows slow decreases (time constant, several hundred milliseconds), which can be partially reversed by flashes. The similarities suggest that the same rate-limiting step governs the opening and closing of channels for both reversible and tethered agonists. Therefore, this step is probably not the initial encounter between agonist and receptor molecules

Constraints on Primordial Nongaussiantiy from the High-Redshift Cluster MS1054--03

The implications of the massive, X-ray selected cluster of galaxies MS1054--03 at $z=0.83$ are discussed in light of the hypothesis that the primordial density fluctuations may be nongaussian. We generalize the Press-Schechter (PS) formalism to the nongaussian case, and calculate the likelihood that a cluster as massive as MS1054 would appear in the EMSS. The probability of finding an MS1054-like cluster depends only on \omegam and the extent of primordial nongaussianity. We quantify the latter by adopting a specific functional form for the PDF, denoted $\psi_\lambda,$ which tends to Gaussianity for $\lambda\gg 1,$ and show how $\lambda$ is related to the more familiar statistic $T,$ the probability of $\ge 3\sigma$ fluctuations for a given PDF relative to a Gaussian. We find that Gaussian initial density fluctuations are consistent with the data on MS1054 only if \omegam\simlt 0.2. For \omegam\ge 0.25 a significant degree of nongaussianity is required, unless the mass of MS1054 has been substantially overestimated by X-ray and weak lensing data. The required amount of nongaussianity is a rapidly increasing function of \omegam for 0.25 \le \omegam \le 0.45, with $\lambda \le 1$ (T \simgt 7) at the upper end of this range. For a fiducial \omegam=0.3, \omegal=0.7 universe, favored by several lines of evidence we obtain an upper limit $\lambda \le 10,$ corresponding to a $T\ge 3.$ This finding is consistent with the conclusions of Koyama, Soda, & Taruya (1999), who applied the generalized PS formalism to low (z\simlt 0.1) and intermediate (z\simlt 0.6) redshift cluster data sets.Comment: 15 pages, 11 figures, submitted to the Astrophysical Journal, uses emulateapj.st

TB10: A Comparison of Arch-yarding and Ground-skidding of Pine Sawlogs in the University of Maine Forest

The decision whether to use a trailing-arch or operate by ground-skidding methods may face many sawlog producers. This case study attempted to determine statistically whether there existed significant differences between the two methods of operation by examining the various activities performed by a crawler-type tractor within the arch-yarding and ground-skidding phases of a harvesting system.https://digitalcommons.library.umaine.edu/aes_techbulletin/1192/thumbnail.jp