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PubMed2002

Presynaptic inhibition of olfactory receptor neurons in crustaceans.

Presynaptic inhibition of transmitter release from primary sensory afferents is a common strategy for regulating sensory input to the arthropod central nervous system. In the olfactory system, presynaptic inhibition of olfactory receptor neurons has been long suspected, but until recently could not be demonstrated directly because of the difficulty in recording from the afferent nerve terminals. A preparation using the isolated but intact brain of the spiny lobster in combination with voltage-sensitive dye staining has allowed stimulus-evoked responses of olfactory receptor axons to be recorded selectively with optical imaging methods. This approach has provided the first direct physiological evidence for presynaptic inhibition of olfactory receptor neurons. As in other arthropod sensory systems, the cellular mechanism underlying presynaptic afferent inhibition appears to be a reduction of action potential amplitude in the axon terminal. In the spiny lobster, two inhibitory transmitters, GABA and histamine, can independently mediate presynaptic inhibition. GABA- and histaminergic interneurons in the lobster olfactory lobe (the target of olfactory receptor neurons) constitute dual, functionally distinct inhibitory pathways that are likely to play different roles in regulating primary olfactory input to the CNS. Presynaptic inhibition in the vertebrate olfactory system is also mediated by dual inhibitory pathways, but via a different cellular mechanism. Thus, it is possible that presynaptic inhibition of primary olfactory afferents evolved independently in vertebrates and invertebrates to fill a common, fundamental role in processing olfactory information.

Animals↗
PubMed2002

Ubiquitin and actin expression in claw muscles of land crab, Gecarcinus lateralis, and American lobster, Homarus americanus: differential expression of ubiquitin in two slow muscle fiber types during molt-induced atrophy.

The closer muscle of large-clawed decapod crustaceans undergoes a proecdysial (premolt) atrophy to facilitate withdrawal of the appendage at ecdysis. This atrophy involves the activation of both calcium-dependent (calpains) and ubiquitin (Ub)/proteasome-dependent proteolytic systems that break down proteins to reduce muscle mass. Moreover, the large slow-twitch (S(1)) fibers undergo a greater atrophy than the small slow-tonic (S(2)) fibers. Both polyUb mRNA and Ub-protein conjugates increase during claw muscle atrophy. In this study in situ hybridization and RT-PCR were used to determine the temporal and spatial expression of polyUb and alpha-actin. A cDNA encoding the complete sequence of lobster muscle alpha-actin was characterized; a probe synthesized from the cDNA provided a positive control for optimizing RT-PCR and in situ hybridization. PolyUb was expressed at low levels in claw closer muscle from anecdysial (intermolt) land crab. By early proecdysis (premolt; stage D(0)), polyUb mRNA levels increased in medial fibers that insert along the midline of the apodeme, with greater expression in S(1) than S(2), while levels remained low in peripheral fibers. By late proecdysis, polyUb mRNA decreased in central fibers, while mRNA increased in peripheral S(1) fibers. In contrast, alpha-actin was expressed in lobster claw muscles at relatively constant levels during the intermolt cycle. These results suggest that Ub/proteasome-dependent proteolysis contributes to enhanced turnover of myofibrillar proteins during claw closer muscle atrophy. Furthermore, atrophy is not synchronous within the muscle; it begins in medial fibers and then progresses peripherally.

Actins↗
PubMed2001

Growth, feeding frequency, protein turnover, and amino acid metabolism in European lobster Homarus gammarus L.

The effect of feeding frequency on growth and protein metabolism in the European lobster, Homarus gammarus, was investigated. Fourth (IV) stage lobsters H. gammarus were fed individually a marine animal meal (herring/mussels meal) for 56 days. Feeding a daily ration equivalent to 10% of their body weight gave better growth than feeding daily rations of 5% and 20%. Protein synthesis rates were similar for the three food rations but protein growth rates were significantly lower and protein degradation rates highest in the 5% body weight per day ration group. The efficiency with which synthesised protein was retained as growth was found to be 38% in the in the 10% ratio group. Protein synthesis rates of lobsters were found to be lower than those for shrimps (Penaeus vannamei). The amino acid flux also suggests a lower protein conversion efficiency than shrimps P. vannamei. The results suggests that lobsters are slow, periodic feeders and that growth can be readily increased by manipulation of particular environmental factors such as feeding frequency.

Amino Acids↗
PubMed2001

Nonuniform distribution of myosin light chains within the thick filaments of lobster slow muscle: Immunocytochemical study.

The in situ distribution of the alpha and beta myosin light chains was investigated at the subsarcomeric and subfilament levels in individual fibers of the superficial flexor muscle (SFM) of the lobster, Homarus americanus. Polyclonal antibodies were produced against the two classes of myosin light chains and used for subsequent immunolocalization on thin sections of sarcomeres and on isolated filaments from both the medial and lateral fiber bundles of the SFM. The beta myosin light chains were uniformly distributed within the crossbridge region of sarcomeres of both medial and lateral bundles. The alpha myosin light chains were uniformly distributed within the crossbridge region of sarcomeres from the medial bundle, but were nonuniformly distributed over the crossbridge region of lateral bundle sarcomeres. In the latter, the number of alpha myosin light chains was highest toward the center of the thick filaments, diminishing towards the ends. Similar distributions of alpha light chains were found in isolated myosin filaments. These data demonstrate that heterogeneity in protein composition extends to the level of the myosin filament and suggest that the myosin filament substructure in lobster may be different than that found in vertebrate skeletal muscle.

Animals↗
PubMed2001

Transport of (22)Na(+) and( 45)Ca(2+) by Xenopus laevis oocytes expressing mRNA from lobster hepatopancreas.

This paper describes the development of a functional assay system to express crustacean epithelial electrogenic 2Na(+)/1H(+) antiporters in Xenopus laevis oocytes. Subsequent publications will use this assay method to establish nucleotide and amino acid sequence information about this transporter by functionally screening an hepatopancreatic cDNA library. In this method, oocytes were injected with hepatopancreatic mRNA (50 ng) isolated from Homarus americanus, while control oocytes received injections of an equivalent volume of distilled water. Three to five days post-injection, oocytes were incubated in media containing either (22)Na(+) or (45)Ca(2+) for specific time intervals and the rates of ion transfer into the oocytes were monitored under a variety of experimental conditions. Uptakes of both radiolabelled cations were stimulated by mRNA injection. mRNA-stimulated (22)Na(+) uptake was significantly (P < 0.05) inhibited by addition of calcium, amiloride, or by an antiporter-specific monoclonal antibody to the external medium. mRNA-stimulated (45)Ca(2+) uptake was significantly (P < 0.05) inhibited by addition of sodium, amiloride, cadmium, zinc, or by the antiporter-specific monoclonal antibody (also inhibitory for (22)Na(+) transport) to the external medium. The kinetics of (22)Na(+) influx in mRNA-injected oocytes were sigmoidal functions of external sodium concentration, exhibiting a Hill Coefficient (n) of approximately 3.0. Both calcium and amiloride significantly (P < 0.05) reduced sigmoidal sodium influx kinetics by alterations in the J(max) (amiloride) or K(Na) (calcium) of the transporter. Size fractionation of hepatopancreatic mRNA resulted in a single fraction that was most stimulatory for sodium and calcium transport and which likely contains the antiporter transcript. The results of this study provide the basis for using (22)Na(+) and (45)Ca(2+) transport assays of lobster mRNA-injected oocytes to functionally screen an hepatopancreatic cDNA library for clones that will provide full length nucleotide and amino acid sequences of the invertebrate electrogenic 2Na(+)/1H(+) antiporter protein.

Amiloride↗
PubMed1975

Comparative strategies in the investigation of neural networks.

Comparative studies on nervous systems, though infrequently undertaken for the purpose of comparison, have yielded some important generalities about the formats of nervous networks, and about the cell biology of certain neural types. In the first category, it is clear that convergent evolutionary processes arrived at very similar networks to accomplish reciprocal and lateral inhibition, and load-compensation in "resistance reflexes." A newer general network format is described, command-derived inhibition, in which the central nervous elements controlling a rapid movement deliver presynaptic inhibition to the terminals of sensory neurons that carry reafferent excitation from the movement. It is argued that such circuits occur in several groups of animals, and that they include as a special class the efferent inhibitory neurons innervating acoustico-lateralis receptors in vertebrates. The properties of circuit elements that now seem to constitute useful generalizations include size principle (the inverse relationship between size and excitability in a variety of neurons), and the late differentiation of sensory neurons, failure to decussate, and their inability to mediate inhibition. Many other generalities have emerged, only to fall; one conclusion from such searches is that many supposedly "basic" properties of cell types or neural circuits are in fact not phylogenetically conservative, however much the physiologist may expect them to be.

Action Potentials↗
PubMed1975

Photoreceptor processes: some problems and perspectives.

Visual photoreceptors from both vertebrates and invertebrates are characterized by extensive elaboration of membrane which contains visual pigment (rhodopsin). Visual pigments in all phyla examined are chemically similar: the chromophore is 11-cis retinaldehyde attached by an aldimine linkage (Schiff base) to a membrane protein, opsin. The effect of light is to isomerize the chromophore to the all-trans configuration. Beyond these fundamental similarities, several specific areas are discussed in which variations and differences appear. (1) Light causes vertebrate visual pigments to bleach, liberating the chromophore. Most invertebrate visual pigments do not bleach in the light, but instead form a thermally stable metarhodopsin, with the chromophore in the all-trans configuration still attached to the opsin. (2) In the disk membranes of vertebrate rod and cone outer segments, the rhodopsin molecules are oriented with their chromophores nearly coplanar with the disks. Within this plane, however, both rotational and translational diffusion are possible. In the microvillar membranes of arthropod and cephalopod rhabdoms, on the other hand, the situation is less clear. There is evidence for some preferential orientation of chromophores that implies restrictions on Brownian rotation. (3) In the outer segments of vertebrate receptors, absorption of light by rhodopsin causes the plasma membrane to hyperpolarize due to a decrease in sodium conductance, possibly mediated by calcium ions. In most invertebrate photoreceptors, light causes a depolarization due to an increase in conductance, principally to sodium ions. A subsequent entry of calcium causes a partial repolarization of the membrane, due to a decrease in sodium conductance. (4) For vertebrate receptors, log threshold is directly proportional to the fraction of rhodopsin bleached (Dowling-Rushton relationship). The proportionality constant varies in different preparations from less than four to more than 30, and the physical basis for the relationship is unknown. For invertebrates, by contrast, the dependence of sensitivity on rhodopsin concentration is much less dramatic and may well depend simply on the probability of quantum catch. (5) In most species, vertebrate and invertebrate, the accumulation of photoproduct probably has no effect on membrane conductance, but several possible exceptions exist. (6) Photoregeneration of rhodopsin from metarhodopsin is likely an important mechanism of recovery in certain arthropods such as diurnal insects, but dark mechanisms of recovery also exist in all phyla. In no single case are they adequately understood.

Adaptation, Ocular↗
PubMed1976

Adaptation of muscle pyruvate kinases to environmental temperatures and pressures.

Kinetic and structural properties of muscle pyruvate kinases from species adapted to different temperatures and hydrostatic pressures were examined. Most muscle pyruvate kinases can exist in two temperature-dependent conformational states. For the low temperature conformers, activation free energies and enthalpies were proportional to the species' adaptation temperatures. The interspecific differences in activation parameters may be due to different amounts of weak bond formation/rupture during the rate-limiting event(s) in catalysis. When comparisons of substrate binding ability at physiological temperatures are made, a marked conservation in apparent Km values is observed among species, Structural rigidity, as estimated by heat inactivation temperature, is positively correlated with adaptation temperature. The apparent size of the conformational change which occurs during catalysis is also positively correlated with cell temperature, except for enzymes for deep-sea species.

Adaptation, Physiological↗
PubMed1981

Synaptic mechanisms that generate network oscillations in the absence of discrete postsynaptic potentials.

Synaptic mechanisms were examined in the pyloric network of the lobster stomatogastric which generate network oscillations in the absence of discrete postsynaptic potentials (PSPs). In normal saline, the unstimulated pyloric network underwent weak bursting in only a few cells. Stimulation of the input nerve, or bath application of the input neurotransmitter dopamine, produced similar vigorous bursting in many pyloric neurons. In saline-containing tetrodotoxin (TTX) plus dopamine, action potentials and corresponding discrete PSPs were blocked, but the underlying slow wave oscillations in network neurons continued. No oscillations occurred in TTX-saline without dopamine. The generation of these nonspiking network oscillations can be explained by the interaction between two synaptic mechanisms which do not produce discrete PSPs: neurotransmitter activation of bursting pacemaker oscillations in a single network neuron, and graded inhibition between network neurons.

Action Potentials↗
PubMed1984

Firing patterns of closer motoneurons during reflex activity in the dimorphic claws of the lobster Homarus americanus.

The in vivo firing patterns of the fast closer excitor (FCE) and slow closer excitor (SCE) motoneurons to the dimorphic claws of the lobster Homarus americanus were analyzed during reflex closure activity by identifying postsynaptic potentials in muscle fibers with known motor innervation. Three types of claw activity were observed: slow closure, rapid closure, and maintained closure. Slow closure and rapid closure in both claws were mediated by SCE and FCE, respectively. In the cutter, maintained closure was mediated only by SCE; in the crusher, both FCE and SCE could maintain closure. The homologous SCEs displayed no significant differences in activity; in both claws, they fired at medium-to-high frequency. The homologous FCEs did display different spike frequencies during claw closure. The crusher FCE fired at high frequencies; the cutter FCE fired at much lower frequencies. Such in vivo differences in axon activity between homologous FCEs are correlated with claw dimorphism and with the population of muscle fiber types.

Animals↗
PubMed1985

Heterogeneity of myofibrillar proteins in lobster fast and slow muscles: variants of troponin, paramyosin, and myosin light chains comprise four distinct protein assemblages.

Fast and slow muscles from the claws and abdomen of the American lobster Homarus americanus were examined for adenosine triphosphatase (ATPase) activity and for differences in myofibrillar proteins. Both myosin and actomyosin ATPase were correlated with fiber composition and contractile speed. Four distinct patterns of myofibrillar proteins observed in sodium dodecyl sulfate-polyacrylamide gels were distinguished by different assemblages of regulatory and contractile protein variants. A total of three species of troponin-T, five species of troponin-I, and three species of troponin-C were observed. Lobster myosins contained two groups of light chains (LC), termed "alpha" and "beta." There were three alpha-LC variants and two beta-LC variants. There were no apparent differences in myosin heavy chain, actin, and tropomyosin. Only paramyosin showed a pattern completely consistent with muscle fiber type: slow fibers contained a species (105 kD) slightly smaller than the principle variant (110 kD) in fast fibers. It is proposed that the type of paramyosin present could provide a biochemical marker to identify the fiber composition of muscles that have not been fully characterized. The diversity of troponin and myosin LC variants suggests that subtle differences in physiological performance exist within the broader categories of fast- and slow-twitch muscles.

Actomyosin↗