Effect of 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) on monoamine neurotransmitters in mouse brain & heart

The effect of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) was studied on dopamine (DA), norepinephrine (NE), serotonin (5HT) and γ-aminobutyric acid (GABA) neurons in mouse brain and on NE neurons of mouse heart. MPTP (45 mg/kg) was administered s.c. to mice twice daily for 2 consecutive days. This dosage regimen produced a decrease in the forebrain concentrations of DA and NE at 7 and 20 days after injection. In contrast, the forebrain concentrations of 5HT and GABA were not significantly decreased at either time. MPTP administration also produced a marked decrease in the uptake of ³H-DA into striatal slices and ³H-NE into cerebral cortical slices. In contrast, the uptake of ³H-NE into hypothalamic slices and the uptake of ³H-5HT into slices from several brain regions were not altered. MPTP initially reduced the concentration of NE in the heart, but unlike the persistent decreases in the forebrain concentrations of NE and DA, the NE concentration in the heart returned to control levels at approximately 20 days after MPTP administration. These results, showing that MPTP can produce a long lasting and selective decrease in the forebrain concentrations of NE and DA and in the uptake of radioactive DA and NE into brain slices, suggest that MPTP can cause the destruction of catecholamine neurons in mouse brain. In contrast, MPTP administration does not appear to produce long term changes in either 5HT or GABA neurons.     

THE UPTAKE OF [3H]GABA BY SLICES OF RAT CEREBRAL CORTEX

—A rapid accumulation of [³H]GABA occurs in slices of rat cerebral cortex incubated at 25° or 37° in a medium containing [³H]GABA. Tissue medium ratios of almost 100:1 are attained after a 60 min incubation at 25°. At the same temperature no labelled metabolites of GABA were found in the tissue or the medium. The process responsible for [³H]GABA uptake has many of the properties of an active transport mechanism: it is temperature sensitive, requires the presence of sodium ions in the external medium, is inhibited by dinitrophenol and ouabain, and shows saturation kinetics. The estimated K_m value for GABA is 2·2 × 10^?5m , and V_max is 0·115 μmoles/min/g cortex. There is only negligible efflux of the accumulated [³H]GABA when cortical slices are exposed to a GABA-free medium. [³H]GABA uptake was not affected by the presence of large molar excesses of glycine, l -glutamic acid, l -aspartic acid, or β-aminobutyrate, but was inhibited in the presence of l -alanine, l -histidine, β-hydroxy-GABA and β-guanidinopropionate. It is suggested that the GABA uptake system may represent a possible mechanism for the inactivation of GABA or some related substance at inhibitory synapses in the cortex.     

Agonistic action of synthetic analogues of quisqualic acid at the insect neuromuscular junction

One hundred twenty analogues of quisqualic acid were synthesized and assayed on the neuromuscular junction of larva of the mealworm, Tenebrio molitor. Two new agonists for amino acid receptors, L-glutamic acid N-thiocarboxyanhydride (L-GANTA) and DL-hydantoinpropionic acid (DL-HPA), were discovered in this study. L-GANTA and DL-HPA produced muscle membrane depolarization, accompanied by a reduction of the muscle input resistance. The amplitude of excitatory postsynaptic potentials was decreased in the presence of L-GANTA and DL-HPA. The apparent dissociation constants obtained from dose-depolarization plots were 7 x 10^?4 M for L-GANTA and 9 x 10^?4 M for DL-HPA. Some structural constraints imposed on agonists at amino acid receptors on insect muscle were discussed.     

γ-Aminobutyric acid-stimulated chloride permeability in crayfish muscle

γ-Aminobutyric acid selectively increased Cl^? permeability in isolated strips of crayfish abdominal muscle. Muscle fibers incubated in VAn Harreveld's solution at room temperature took up ³⁶Cl^? to the extent of 700 ml/kg wet weight with a halftime of 2.5 min. During 15-s incubations, the control ³⁶Cl^? uptake space was 131 ± 4 ml/kg (n = 60) and this was significantly increased by γ-aminobutyric acid at 200 μM or higher concentrations to 177 ± 4 ml/kg (n = 48, P < 0.05). This effect was specific for chloride since γ-aminobutyric acid did not increase the uptake by crayfish muscle of radioactive sucrose, inositol, or propionate. γ-Aminobutyric acid stimulation of ³⁶Cl^? uptake is mediated by receptor-ionophore function since the process shows pharmacological properties virtually identical to those observed by electrophysiological techniques. The γ-aminobutyric acid stimulation of Cl^? permeability is dose dependent with 50% of the maximal effect at 40 μM γ-aminobutyric acid and the dose vs. response curve is somewhat sigmoid. The γ-aminobutyric acid agonist muscimol causes the same maximal effect on Cl^? uptake as γ-aminobutyric acid, but acts at 5-fold lower concentrations, i.e. is more potent. However, the partial agonist γ-amino, β-hydroxybutyric acid produced little or no stimulation of ³⁶Cl^? flux. The response to γ-aminobutyric acid was blocked by 2 mM β-guanidinopropionate or γ-guanidinobutyrate, 0.5 mM bicuculline, and 10 μM picrotoxinin. Picrotoxinin inhibition was dose dependent with 50% inhibition occurring at 4 μM. Antagonists did not affect control ³⁶Cl^? uptake. These results confirm electrophysiological observations that the postsynaptic response to the inhibitory neurotransmitter γ-aminobutyric acid involves a rapid increase in membrane permeability to Cl^?     

The metabolism of gamma-aminobutyric acid (GABA) in the lobster nervous system--uptake of GABA in the nerve-muscle preparations

The lack of information on the mechanism of inactivation of the crustacean neuromuscular inhibitory transmitter compound prompted a study of the disposition of radioactive γ-aminobutyric acid (GABA) in lobster nerve-muscle preparations. A specific GABA transport system was found. Radioactive GABA was concentrated by the tissues to levels several times those in the medium, and net uptake could be demonstrated. The process was dependent on sodium ions in the medium; neither lithium nor choline could substitute for sodium. Incubations with increasing GABA concentrations indicated that uptake was a saturable mechanism with an apparent K_m of 5.8 × 10⁻⁵m . Of many compounds tested, only desmethylimipramine, chlopromazine (and several related compounds), and certain close structural analogues (guanidinoacetic acid, β-guani-dinopropionic acid and,β-hydroxy-GAB A) were effective inhibitors of uptake. The inhibition with all these compounds, however, was at high concentrations (5 × 10⁻⁴ to 10⁻³m ) which limited their usefulness for physiological studies. A separate uptake mechanism for glutamate was found in the lobster nerve-muscle preparations. This process was not described in detail, but certain properties are similar to those of the GABA transport system. The cellular location of the GABA uptake system remains unknown. By analogy with noradrenaline inactivation, however, it is postulated that uptake could serve to terminate the physiological actions of GABA by rapidly removing it from its sites of action in synaptic clefts.     

Hyperuricemia enhances intracellular urate accumulation via down-regulation of cell-surface BCRP/ABCG2 expression in vascular endothelial cells

Hyperuricemia has been recognized as an independent risk factor for cardiovascular disease. Urate stimulates NADPH oxidase and induces production of reactive oxygen species (ROS); consequently, intracellular urate accumulation can induce oxidative stress leading to endothelial dysfunction. Here, we studied the mechanism involved, using human umbilical vascular endothelial cells (HUVEC) as a model. Pretreatment with 15 mg/dL unlabeled uric acid (corresponding to hyperuricemia) resulted in increased uptake of [¹⁴C]uric acid at steady-state by HUVEC, whereas pretreatment with 5 mg/dL uric acid (in the normal serum concentration range) did not. However, the initial uptake rate of [¹⁴C]uric acid was not affected by uric acid at either concentration. These results suggest that efflux transport of uric acid is decreased under hyperuricemic conditions. We observed a concomitant decrease of phosphorylated endothelial nitric oxide synthase. Plasma membrane expression of breast cancer resistance protein (BCRP), a uric acid efflux transporter, was decreased under hyperuricemia, though the total cellular expression of BCRP remained constant. Uric acid did not affect expression of another uric acid efflux transporter, multidrug resistance associated protein 4 (MRP4). Moreover, phosphorylation of Akt, which regulates plasma membrane localization of BCRP, was decreased. These uric acid-induced changes of BCRP and Akt were reversed in the presence of the antioxidant N-acetylcysteine. These results suggest that in hyperuricemia, uric acid-induced ROS generation inhibits Akt phosphorylation, causing a decrease in plasma membrane localization of BCRP, and the resulting decrease of BCRP-mediated efflux leads to increased uric acid accumulation and dysregulation of endothelial function.     

THE EFFECT OF DELTA-AMINOLAEVULINIC ACID ON THE UPTAKE AND EFFLUX OF [3H]GABA IN RAT BRAIN SYNAPTOSOMES

§-Aminolaevulinic acid (§-ALA) is an omega amino acid which can be considered as an analogue of γ-aminobutyric acid (GABA). We have examined the effect of §-ALA on [³H]GABA uptake and release in the synaptosome fraction of rat cerebral cortex and report: (1) High concentrations of §-ALA (0.75-5 mM) stimulated [³H]GABA release very markedly, the stimulation with 1mM and 5mM-§-ALA exceeding the maximum obtainable with unlabelled GABA; (2) Low concentrations of §-ALA (0.1-0.5 mM) produced little stimulation of [³H]GABA efflux, less than that produced by similar concentrations of unlabelled GABA; (3) 0.1 mM-§-ALA reduced the stimulation of [³H]GABA efflux elicited by 55 mM-K⁺ and the combination of 1 mM-§-ALA and 55mM-K⁺ produced a lower stimulation of efflux than 1 mM-§-ALA alone; (4) §-ALA inhibits [³H]GABA uptake in a linearly competitive fashion and inhibition is maximal at 0.5 mM-§-ALA. These results are discussed in relation to the neuronal high affinity GABA transport mechanism and inhibition of the synaptosomal Na⁺ and K⁺ -dependent ATPase. It is also postulated that §-ALA increases the chloride conductance of the synaptosomal membrane, possibly by acting on presynaptic GABA receptors.     

Effects of adenosine 3′ : 5′-monophosphate and guanosine 3′ : 5′-monophosphate on calcium uptake and phosphorylation in membrane fractions of vascular smooth muscle

The effects of adenosine 3′ : 5′-monophosphate (cyclic AMP), guanosine 3′ : 5′-monophosphate (cyclic GMP) and exogenous protein kinase on Ca uptake and membrane phosphorylation were studied in subcellular fractions of vascular smooth muscle from rabbit aorta. Two functionally distinct fractions were separated on a continuous sucrose gradient: a light fraction enriched in endoplasmic reticulum (fraction E) and a heavier fraction containing mainly plasma membranes (fraction P).While cyclic AMP and cyclic GMP had no effect on Ca uptake in the absence of oxalate, both cyclic nucleotides inhibited the rate of oxalate-activated Ca uptake when used at concentrations higher than 10^?5 M. The addition of bovine heart protein kinase to either fraction produced an increase in the rate of oxalate-activated Ca uptake which was further augmented by cyclic AMP. Cyclic GMP caused smaller stimulations of protein kinase-catalyzed Ca uptake than cyclic AMP.Mg-dependent phosphorylation, attributable to endogenous protein kinase(s), was inhibited in fraction E by low concentrations (10^?8 M) of both cyclic AMP and cyclic GMP. In fraction P, an inhibition by cyclic AMP occurred also at a concentration of 10^?8 M, while with cyclic AMP a concentration of 10^?5 M was required for a similar inhibition. Bovine heart protein kinase stimulated the phosphorylation of the membrane fractions much more than Ca uptake. In fraction E, in the presence of bovine protein kinase, both cyclic AMP and cyclic GMP stimulated phosphorylation up to 200%. Under these conditions, no stimulation was observed in fraction P.These results are compatible with the hypothesis that in vascular smooth muscle soluble rather than particulate protein kinases are involved in the regulation of intracellular Ca concentration.     

Administration of the antibacterial agents flumequine and oxolinic acid to small turbot (Scophthalmusmaximus L.) by bath

Administration of flumequine and oxolinic acid to turbot, Scophthalmus maximus L., by bath resulted in significant levels of both drugs in the muscle tissue. Bath treatment using 150 mg L^?1 of flumequine and 200 mg L^?1 of oxolinic acid for 72 h gave muscle concentrations of 10.2 and 6.2 μg g^?1, respectively. Excretion of both antibacterials was rapid, reaching concentrations of 0.8 and 0.9 μg g^?1, respectively, for flumequine and oxolinic acid 24 h after the end of treatment. At day 3 post‐treatment the concentration of flumequine was below the limit of quantitation (0.1 μg g^?1) of the analytical method. Based on a minimum inhibitory concentration (MIC) of 0.0625 μg ml^?1 for susceptible strains, bath treatment maintain muscle levels in excess of 0.5 μg ml^?1, corresponding to eight times the MIC‐value for approximately 118 h for oxolinic acid and 104 h for flumequine.     

Isoguvacine Binding, Uptake, and Release: Relation to the GABA System

Isoguvacine (1,2,3,6-tetrahydropyridine-4-car-boxylic acid) is a GABA (γ-aminobutyric acid) agonist with limited conformational flexibility. In these studies we investigated the binding, uptake, and release of [³H] isoguvacine by use of tissue preparations of rat CNS, comparing the results with similar studies of [³H]GABA. The results from these investigations indicate that isoguvacine binds to membrane preparations of rat forebrain with pharmacological characteristics similar to the post-synaptic GABA recognition site; that it is transported into synaptosomal preparations by an uptake system similar to the high-affinity GABA uptake system; and that recently accumulated isoguvacine is released in a Ca²⁺-dependent manner and by heteroexchange with external GABA. The ability of isoguvacine and γ-hydroxybutyric acid to decrease the K⁺-stimulated Ca²⁺-dependent release process was also investigated. The results indicate that isoguvacine interactions have many of the biochemical features of GABA synaptic function, isoguvacine being, however, less potent than GABA.     

The electrophysiology and pharmacology of lobster neuromuscular synapses

Effects of drugs on resting potential, membrane resistance, and excitatory and inhibitory postsynaptic potentials (e.p.s.p.'s and i.p.s.p.'s) of lobster muscle fibers were studied using intracellular microelectrodes Acetylcholine, d-tubocurarine, strychnine, and other drugs of respectively related actions on vertebrate synapses were without effects even in 1 per cent solutions (10^- w/v). Gamma-aminobutyric acid (GABA) acted powerfully and nearly maximally at 10^-7 to 10^-6 w/v. Membrane resistance fell two- to tenfold, the resting potential usually increasing slightly. This combination of effects, which indicates activation of inhibitory synaptic membrane, was also produced by other short chain ω-amino acids and related compounds that inactivate depolarizing axodendritic synapses of cat. The conductance change, involving increased permeability to Cl^-, by its clamping action on membrane potential shortened as well as decreased individual e.p.s.p.'s. Picrotoxin in low concentration (ca. 10^-7 w/v) and guanidine in higher (ca. 10^-3 w/v) specifically inactivate inhibitory synapses. GABA and picrotoxin are competitive antagonists. The longer chain ω-amino acids which inactivate hyperpolarizing axodendritic synapses of cat are without effect on lobster neuromuscular synapse. However, one member of this group, carnitine (β-OH-GABA betaine), activated the excitatory synapses, a decreased membrane resistance being associated with depolarzation. The pharmacological properties of lobster neuromuscular synapses and probably also of other crustacean inhibitory synapses appear to stand in a doubly inverted relation to axodendritic synapses of cat.     

Characterization of the membrane bound Mg2+-ATPase of rat skeletal muscle

A procedure was developed to isolate a membrane fraction of rat skeletal muscle which contains a highly active Mg²⁺-ATPase (5–25 μmol P_i/mg min). The rate of ATP hydrolysis by the Mg²⁺-ATPase was nonlinear but decayed exponentially (first-order rate constant ≥0.2 s^?1 at 37°C). The rapid decline in the ATPase activity depended on the presence of ATP or its nonhydrolyzable analog 5′-adenylyl imidodiphosphate (AdoPP[NH]P). Once inactivated, removal of ATP from the medium did not immediately restore the original activity. ATP- or AdoPP[NH]P-dependent inactivation could be blocked by concanavalin A, wheat germ agglutinin or rabbit antiserum against the membrane. Additions of these proteins after ATP addition prevented further inactivation but did not restore the original activity. Low concentrations of ionic and nonionic detergents increased the rate of ATP-dependent inactivation. Higher concentrations of detergents, which solubilize the membrane completely, inactivated the Mg²⁺-ATPase. Cross-linking the membrane components with glutaraldehyde prevented ATP-dependent inactivation and decreased the sensitivity of the Mg²⁺-ATPase to detergents. It is proposed that the regulation of the Mg²⁺-ATPase by ATP requires the mobility of proteins within the membrane. Cross-linking the membrane proteins with lectins, antiserum or glutaraldehyde prevents inactivation; increasing the mobility with detergents accelerates ATP-dependent inactivation.     

13C Metabolic Flux Analysis of Escherichia coli Engineered for Gamma‐Aminobutyrate Production

Escherichia coli is engineered for γ‐aminobutyrate (GABA) production in glucose minimal medium. For this, overexpression of mutant glutamate decarboxylase (GadB) and mutant glutamate/GABA antiporter (GadC), as well as deletion of GABA transaminase (GabT), are accomplished. In addition, the carbon flux to the tricarboxylic acid cycle is engineered by the overexpression of gltA, ppc, or both. The overexpression of citrate synthase (CS), encoded by gltA, increases GABA productivity, as expected. Meanwhile, the overexpression of phosphoenolpyruvate carboxylase (PPC) causes a decrease in the rate of glucose uptake, resulting in a decrease in GABA production. The phenotypes of the strains are characterized by ¹³C metabolic flux analysis (¹³C MFA). The results reveal that CS overexpression increases glycolysis and anaplerotic reaction rates, as well as the citrate synthesis rate, while PPC overexpression causes little changes in metabolic fluxes, but reduces glucose uptake rate. The engineered strain produces 1.2 g L^?1 of GABA from glucose. Thus, by using ¹³C MFA, important information is obtained for designing metabolically engineered strains for efficient GABA production.     

The evolution of insulin resistance in muscle of the glucose infused rat

Glucose infusion into rats causes skeletal muscle insulin resistance that initially occurs without changes in insulin signaling. The aim of the current study was to prolong glucose infusion and evaluate other events associated with the transition to muscle insulin resistance. Hyperglycemia was produced in rats by glucose infusion for 3, 5 and 8 h. The rate of infusion required to maintain hyperglycemia was reduced at 5 and 8 h. Glucose uptake into red quadriceps (RQ) and its incorporation into glycogen decreased between 3 and 5 h, further decreasing at 8 h. The earliest observed change in RQ was decreased AMPKα2 activity associated with large increases in muscle glycogen content at 3 h. Activation of the mTOR pathway occurred at 5 h. Akt phosphorylation (Ser⁴⁷³) was decreased at 8 h compared to 3 and 5, although no decrease in phosphorylation of downstream GSK-3β (Ser⁹) and AS160 (Thr⁶⁴²) was observed. White quadriceps showed a similar but delayed pattern, with insulin resistance developing by 8 h and decreased AMPKα2 activity at 5 h. These results indicate that, in the presence of a nutrient overload, alterations in muscle insulin signaling occur, but after insulin resistance develops and appropriate changes in energy/nutrient sensing pathways occur.     

Oxidative Metabolism of 4-Aminobutyrate by Rat Brain Mitochondria: Inhibition by Branched-chain Fatty Acid

Abstract: The oxidation of 4-aminobutyric acid (GABA) by nonsynaptosomal mitochondria isolated from rat forebrain and the inhibition of this metabolism by the branched-chain fatty acids 2-methyl-2-ethyl caproate (MEC) and 2, 2-dimethyl valerate (DMV) were studied. The rate of GABA oxidation, as measured by O₂ uptake, was determined in medium containing either 5 or 100 mM-[K⁺]. The apparent K_m for GABA was 1.16 ± 0.19 mM and the V_max in state 3 was 23.8 ± 5.5 ng-atoms O₂. min^?1. mg protein^?1 in 5 mM-[K⁺]. In a medium with 100 mM-[K⁺] the apparent K_m was 1.11 ± 0.17 mM and V_max was 47.4 ± 5.7 ng-atoms O₂. min^?1. mg protein^?1. The K_m for MEC was determined to be 0.58 ± 0.24 or 0.32 ± 0.08 mM, in 5 or 100 mM-[K⁺], respectively. For DMV, the K_i was 0.28 ± 0.05 or 0.34 ± 0.06 mM, in 5 or 100 mM-[K⁺] medium, respectively. The O₂ uptake of the mitochondria in the presence of GABA was coupled to the formation of glutamate and aspartate; the ratio of oxygen uptake to the rate of amino acid formation was close to the theoretical value of 3. Neither the [K²] nor any of the above inhibitors had any effect on this ratio. The metabolism of exogenous succinic semialdehyde (SSA) by these same mitochondria was also examined. The V_max for utilization of oxygen in the presence of SSA was much greater than that found with exogenously added GABA, indicating that the capacity for GABA oxidation by these mitochondria is not limited by SSA dehydrogenase. In addition, the branched-chain fatty acids did not inhibit the metabolism of exogenously added SSA. Thus, the inhibitors examined apparently act by competitively inhibiting the GABA transaminase system of the mitochondria.     

Elevated CO2 plus chronic warming reduce nitrogen uptake and levels or activities of nitrogen‐uptake and ‐assimilatory proteins in tomato roots

Atmospheric CO₂ enrichment is expected to often benefit plant growth, despite causing global warming and nitrogen (N) dilution in plants. Most plants primarily procure N as inorganic nitrate (NO₃^?) or ammonium (NH₄⁺), using membrane‐localized transport proteins in roots, which are key targets for improving N use. Although interactive effects of elevated CO₂, chronic warming and N form on N relations are expected, these have not been studied. In this study, tomato (Solanum lycopersicum) plants were grown at two levels of CO₂ (400 or 700 ppm) and two temperature regimes (30 or 37°C), with NO₃^? or NH₄⁺ as the N source. Elevated CO₂ plus chronic warming severely inhibited plant growth, regardless of N form, while individually they had smaller effects on growth. Although %N in roots was similar among all treatments, elevated CO₂ plus warming decreased (1) N‐uptake rate by roots, (2) total protein concentration in roots, indicating an inhibition of N assimilation and (3) shoot %N, indicating a potential inhibition of N translocation from roots to shoots. Under elevated CO₂ plus warming, reduced NO₃^?‐uptake rate per g root was correlated with a decrease in the concentration of NO₃^?‐uptake proteins per g root, reduced NH₄⁺ uptake was correlated with decreased activity of NH₄⁺‐uptake proteins and reduced N assimilation was correlated with decreased concentration of N‐assimilatory proteins. These results indicate that elevated CO₂ and chronic warming can act synergistically to decrease plant N uptake and assimilation; hence, future global warming may decrease both plant growth and food quality (%N).     

Comparison of synaptosomal and glial uptake of pipecolic acid and GABA in rat brain

The active uptake of [³H]pipecolic acid increased with incubation time and its uptake at 3 min was half of that at 20 min. [¹⁴C]GABA uptake rose earlier, and its uptake at 3 min was almost 80% of that at 20 min. On the other hand, a ratio (pellet/medium) of [³H]pipecolic acid uptake into glial cell-enriched fractions, was much less (0.4–0.6) than that of [¹⁴C]GABA (25.8–74.1). GABA, 10^–4 M, and pipecolic acid, 10^–4 M, produced a significant inhibition of [³H]pipecolic acid uptake into P₂ fractions. Pipecolic acid, 10^–4 M, significantly reduced the synaptosomal and glial uptake of [¹⁴C]GABA. GABA, 10^–4 M, affected neither spontaneous nor high K⁺-induced release of [³H]pipecolic acid from brain slices. It is suggested that pipecolic acid is involved in either synaptic transmission or in its modulation at GABA synapses in the central nervous system.     

Neuromuscular modulation in Limulus by both octopamine and proctolin

Both octopamine and proctolin potentiate nerve-evoked skeletal muscle contractions in the horseshoe crab, Limulus. The threshold concentration for octopamine was 10^?9 to 10^?8M, while for proctolin it was 3 × 10^?9M. Norepinephrine and dopamine produced effects similar to octopamine but at higher thresholds; tyramine and serotonin were ineffective. Octopamine caused significant increases in amplitudes of excitatory postsynaptic potentials (epsps) of muscle fibers, but had little effect on muscle fiber input resistance or membrane potential. Also, octopamine did not affect depolarization of muscle fibers and subsequent contraction due to the direct action of exogenously applied glutamate. These results suggest that octopamine potentiates nerve-evoked contractions primarily by facilitating release of neuromuscular transmitter. At concentrations above 10^?7M, however, octopamine sometimes caused muscle spikes in response to motoneuron stimulation, a finding that suggests that octopamine may also have some postsynaptic action. Proctolin potentiated the muscle contractions evoked by glutamate but had little effect on glutamate-evoked muscle fiber depolarization, muscle fiber input resistance, or membrane potential. Thus, proctolin appears to act directly on skeletal muscle to enhance contractility. The proctolin-induced potentiations of contraction were sometimes accompanied by modest increases in epsp amplitude, so that unlike lobster skeletal and Limulus cardiac neuromuscular preparations, proctolin may have a secondary direct synaptic effect. Both octopamine and proctolin have been found in Limulus cardiac ganglion. This potential access to the hemolymph and the relatively low threshold concentrations needed for physiological action suggest that octopamine and proctolin could function as hormonal modulators of neuromuscular function in Limulus.     

A Calcium Aspect of the Effect of Noradrenaline on the Resting Membrane Potential in Somatic Muscle Cells of the Earthworm

A hyperpolarizing effect of noradrenaline (NA) on muscle cells of the earthworm caused by activation of the membrane ion pumps is eliminated in a Ca-free medium, in the case of replacement of Na⁺ by Mn²⁺, or when verapamil or chlorpromazine have been added to the bath solution. A decrease or an increase in the Ca²⁺ concentration in the solution, as well as caffeine application, do not influence the resting membrane potential (RMP) of muscle cells. It is supposed that signal transmission from the membrane adrenoreceptors to the ion pump of earthworm muscle cells by NA is provided via entry of extracellular Ca²⁺ ions into the cell with subsequent involving of Ca²⁺ acceptor proteins similar to calmodulin in vertebrate animals.     

δ-Aminolaevulinic Acid Uptake, Toxicity, and Effect on [14C]γ-Aminobutyric Acid Uptake into Neurons and Glia in Culture

δ-Aminolaevulinic acid (ALA) uptake into neurons and glia in primary culture as well as ALA toxicity and its effects on γ-aminobutyric acid (GABA) uptake were examined. [4-¹⁴C]ALA uptake into neurons and glia was nonsaturable, partially Na⁺- and temperature-dependent, and appeared to comprise mainly diffusion into the cell. 2,4-Dinitrophenol caused some inhibition of [4-¹⁴C]ALA uptake whereas ouabain, KCN, or amino acids at 1 mM concentration were without effect. ALA (1 mM) caused a slight inhibition of [U-¹⁴C]GABA uptake into neurons (14%) and glia (9%), but was without effect at lower concentrations. It is unlikely that, in acute porphyria, ALA reaches sufficiently high levels in nervous tissue to interfere with the reuptake of GABA into neurons or glia. ALA was shown to be toxic, judged by the loss of cells, to both neurons and glia at concentrations as low as 10 μM. Such a concentration of ALA may be expected to occur in the CSF of porphyric patients in the acute attack. However, results obtained with dispersed cells in culture may not necessarily reflect the situation in vivo where the cell may have a far greater resistance to the effects of toxic agents.