Fast Closer Excitor Research Articles

The Neuromuscular Junctions of the Slow and the Fast Excitatory Axon in the Closer of the CrabEriphia spinifronsAre Endowed with Different Ca2+Channel Types and Allow Neuron-Specific Modulation of Transmitter Release by Two Neuropeptides

Most crustacean muscle fibers receive double excitatory innervation by functionally different motor neurons termed slow and fast. By using specific omega-toxins we show that the terminals of the slow closer excitor (SCE) and the fast closer excitor (FCE) at a crab muscle are endowed with different sets of presynaptic Ca(2+) channel types. omega-Agatoxin, a blocker of vertebrate P/Q-type channels, reduced the amplitude of EPSCs by decreasing the mean quantal content of transmitter release in both neurons by 70-85%, depending on the concentration. We provide the first evidence that omega-conotoxin-sensitive channels also participate in transmission at crustacean neuromuscular terminals and are colocalized with omega-agatoxin-sensitive channels in an axon-type-specific distribution. omega-Conotoxin, a blocker of vertebrate N-type channels, inhibited release by 20-25% only at FCE, not at SCE endings. Low concentrations of Ni(2+), which block vertebrate R-type channels, inhibited release in endings of the SCE by up to 35%, but had little effects in FCE endings. We found that two neuropeptides, the FMRFamide-like DF(2) and proctolin, which occur in many crustaceans, potentiated evoked transmitter release differentially. Proctolin increased release at SCE and FCE endings, and DF(2) increased release only at FCE endings. Selective blocking of Ca(2+) channels by different omega-toxins in the presence of peptides revealed that the target of proctolin-mediated modulation is the omega-agatoxin-sensitive channel (P/Q-like), that of DF(2) the omega-conotoxin-sensitive channel (N-like). The differential effects of these two peptides allows fine tuning of transmitter release at two functionally different motor neurons innervating the same muscle.

Presynaptic inhibition and the participation of GABA(B) receptors at neuromuscular junctions of the crab Eriphia spinifrons.

Presynaptic inhibition exerted by the common inhibitor on the closer and opener muscles and by the specific inhibitor on the opener muscle was investigated in the crab Eriphia spinifrons. In the closer muscle, activation of GABA(B) receptors by baclofen reduced the mean quantal content of excitatory junctional currents by about 25%. Blocking GABA(B) receptors with CGP 55845 diminished presynaptic inhibition at a similar percentage. GABA(B) receptor-mediated presynaptic inhibition is linked to G proteins. Application of pertussis toxin eliminated about 25% of the inhibition exerted by the common inhibitory neuron. GABA(B) receptors participate in presynaptic inhibition at release boutons of the slow and the fast closer excitor at a similar percentage. In the opener muscle, presynaptic inhibition of transmitter release from the same endings of the opener excitor was about 15% stronger with the specific inhibitor than with the common inhibitor. About 10% of the presynaptic inhibition produced by either one of the two inhibitors could be abolished by blocking GABA(B) receptors. The amplitudes of the excitatory junctional currents in the opener were reduced in the presence of baclofen by about 25%, suggesting that synaptic terminals of the opener excitor are endowed with a similar percentage of GABA(B) receptors as terminals of the slow and the fast closer excitors. Baclofen had no effect on postsynaptic inhibition, indicating that GABA(B) receptors are not involved in postsynaptic neuromuscular inhibition.

Allatostatin modulates skeletal muscle performance in crustaceans through pre- and postsynaptic effects.

Allatostatins, originally identified in insects as peptide inhibitors of juvenile hormone biosynthesis, are regarded as potent inhibitory regulators of intestinal muscles in insects and crustaceans. However, accumulating data indicate that allatostatins might also be involved in modulation of skeletal neuromuscular events. We show that most ganglia of two isopod crustaceans (Idotea baltica and I. emarginata) contain pairs of large, allatostatin-immunoreactive motor neurons which supply several segmental muscles. Among them are the dorsal extensor muscles, of which some fibres receive immunoreactive, varicose innervation. We demonstrate, on identified muscle fibres, that allatostatin exerts a twofold inhibitory effect: it reduces contractions of single voltage-clamped fibres, and it decreases the amplitude of evoked excitatory junctional currents recorded from individual release boutons. No change in excitation-contraction threshold or in passive membrane parameters was observed. As the amplitude of miniature currents generated by spontaneously released single transmitter quanta was not changed, the inhibitory effect of the peptide on junctional currents must be of presynaptic origin. Supportive results were obtained on leg muscles of the crab Eriphia spinifrons, where allatostatin decreased evoked synaptic currents by reducing the mean number of transmitter quanta released by presynaptic depolarization without affecting the amplitudes of currents generated by single quanta. This effect of allatostatin was similar for two functionally different neurons, the slow and the fast closer excitor. The data show that allatostatin occurs in identified motor neurons of Idotea and exerts complementary pre- and postsynaptic modulatory effects which reduce muscle responses. Thus, allatostatin counteracts the effects of another neuropeptide, proctolin, which is also present in Idotea and causes potentiating effects on the same muscle fibres.

Fibre Heterogeneity in the Closer and Opener Muscles of Crayfish Walking Legs

ABSTRACT The closer and opener muscles in the third walking legs of the three crayfish Pacifastacus leniusculus, Procambarus clarkii and Astacus leptodactylus are composed of fibres which differ in histochemistry, electrophysiology and morphology. Three major groups of fibres (A, B and C) were distinguished. Group A fibres react weakly to histochemical stains for myofibrillar ATPase (mATPase) activity characteristic of fibres with slow shortening speeds. In the opener muscle, they are innervated by the opener excitor (OE) and the specific opener inhibitor (OI). In the closer muscle, group A fibres are innervated by the common inhibitory neurone (CI) in addition to single (slow closer excitor, SCE) or double excitatory (SCE and fast closer excitor, FCE) innervation. Group A fibres have the largest excitatory junction potentials (EJPs), the longest membrane time constants (τ) and the longest sarcomeres. They are located at the very distal and proximal ends of both muscles. Group B fibres show higher mATPase activity than group A fibres. In the opener muscle, they are innervated by OE and OI; in the closer muscle, they receive double excitatory (SCE and FCE) and CI innervation. Single SCE and OE EJPs are small; those caused by FCE are larger. τ is shorter than in the other two fibre groups. Sarcomere lengths lie between those of group A and C fibres. Group B fibres are found along the entire lengths of both muscles. Group C fibres exhibit the highest mATPase activity (characteristic of fibres with fast shortening velocity) which, in contrast to the ATPase of group B fibres, is not resistant to alkaline preincubation at pH10.05. In the closer, these fibres lack innervation by CI, otherwise the innervation pattern is identical to that of group B fibres. EJP size is similar to that of group B fibres; τ ranges between values for group A and B fibres. Sarcomere lengths are the shortest of all the fibre types. Group C fibres constitute the majority of the fibres in the two muscles and mainly occupy the central regions.

Synaptic plasticity in a regenerated crayfish phasic motoneuron.

Crustacean neuromuscular systems provide many advantages for the study of synaptic transmission and plasticity. The present study examines aspects of synaptic transmission in the phasic, fast closer excitor (FCE) motoneuron of regenerated crayfish claws. Excitatory postsynaptic potentials (EPSPs) fatigued rapidly and showed poor long-term facilitation (LTF) in the smallest of regenerating claws. EPSPs in larger regenerating claws fatigued less and showed pronounced facilitation. These observations were not the same as those previously made during primary development of this motoneuron (Lnenicka and Atwood, 1985a, J. Neuroscience 5:459-467). Hence, regeneration is not the recapitulation of primary development. In situ stimulation of the FCE is known to lead to long-lasting adaptation of synaptic performance. This adaptation is age dependent; it is expressed in young but not old animals. In the regenerated FCE of old animals, we observed a novel form of long-lasting adaptation to imposed activity: EPSPs showed large initial EPSPs and did not exhibit resistance to fatigue during maintained stimulation. This indicates that aged motoneurons can express adaptive changes to increased activity following axonal regeneration, but that the adaptive changes are the opposite to what is observed in nonregenerated motoneurons.

Early experience influences the development of bilateral asymmetry in a lobster motoneuron

The development of functional asymmetry between a pair of homologous motoneurons of the claw closer muscles in lobsters, Homarus americanus, was studied. In juvenile lobsters, 3–5 years old, where the paired claws are highly specialized into a major (crusher) and minor (cutter) type, the fast closer excitor (FCE) motoneuron fired longer bursts of spikes in the crusher claw compared to those in its cutter counterpart. The intraburst impulse frequency was greater for the cutter FCE and its neuromuscular synapses showed greater facilitation at these high impulse frequencies compared to that of the crusher claw. However, such asymmetry in firing patterns and synaptic facilitation was absent in lobsters raised without a substrate and having paired cutter claws. In the earliest juvenile stage, synaptic facilitation was similar between the paired claws and then developed in either an asymmetric or symmetric manner depending on whether the lobsters experienced a substrate or not. In a substrate-free environment asymmetry could be produced by exercising one of the claws during development, implicating bilateral differences in the reflexive activity of the claws as a control mechanism.

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.

Neuromuscular relationships during claw regeneration in Californian snapping shrimp.

We have examined the innervation patterns of the two excitor axons to the closer muscle in the dimorphic (snapper and pincer) claws of Californian snapping shrimp (Alpheus californiensis). In both claws the fast-closer excitor (FCE) axon supplies all of the closer muscle fibers. The slow-closer excitor (SCE) axon, however, makes functional connections only with fibers on the dorsal and ventral margins, which are composed of slow-type fibers in the large snapper claw and of intermediate-type fibers in the smaller pincer claw. The central band of fast fibers in the pincer closer muscle is not innervated by the SCE. During claw regeneration, the closer muscle is initially composed of a population of fast fibers in the pincer and intermediate-type fibers in the snapper. The innervation patterns of the two excitatory motor axons in regenerating claws at this stage are the same as in fully developed claws. During the first intermolt period some fast-closer muscle fibers in the pincer claw differentiate into intermediate-type fibers, but the axon innervation patterns do not change. If the correlation between SCE axon innervation pattern and the regional distribution of different muscle fiber types is indicative of nerve-muscle interactions, the present data suggest that the trophic influence must proceed from nerve to muscle.

Paired impulses in lobster claw motoneurons: in vitro and in vivo production

The somata of the claw opener excitor (OE), the slow closer excitor (SCE), and the fast closer excitor (FCE) motoneurons were physiologically identified in the lobster, Homarus americanus, using an isolated claw ganglion preparation. Depolarization of the soma produced paired impulses with an interpulse interval of 10–12 ms, which demonstrates that doublet production is an intrinsic motoneuron property. Similar paired impulses were found for the FCE axon in intact lobsters where they occurred with a shorter interpulse interval for the cutter than for the crusher claw.

Development of excitatory innervation in the lobster claw closer muscle.

The development of excitatory innervation to the claw closer muscles of 1st (larval), 4th (juvenile), and adult stage lobster was examined by thin serial section electron microscopy. This was possible since neuromuscular terminals of the excitatory axon are distinguishable from those of the inhibitory axon by the shape of their synaptic vesicles in all three stages. In the adult cutter claw closer muscle, innervation of a dorsal and a ventral fiber which is supplied by the fast closer excitor (FCE) and slow closer excitor (SCE) axons respectively, was qualitatively as well as quantitatively similar. Consequently no attempt was made to distinguish between innervation by FCE and SCE axons in the subsequent analysis of 1st, 4th, and adult stage claw closer muscles. Excitatory innervation in the 1st larval stage is limited to four discrete locations in the entire cross-sectional area of the closer muscle. It subsequently spreads to each individual muscle fiber in the adult, thus demonstrating the tremendous proliferation of innervation during development. Concomitantly the mean size of synapses increases significantly from the 1st stage to the 4th stage to the adult lobster. This increase in synaptic size may occur by both the fusion and enlargement of existing smaller synapses. In contrast, the mean size of presynaptic dense bars and their mean number per synapse remained fairly constant in each of the 1st, 4th, and adult stages. However, a relatively greater proportion of adult synapses possessed two or more dense bars compared to their larval and juvenile counterparts. Development of innervation of the lobster claw closer muscle therefore consists of a substantial proliferation of axonal tissue and enlargement of synaptic size.

Electrical and structural properties of crayfish claw motoneurons in an isolated claw-ganglion preparation

1. An isolated claw-ganglion preparation of the crayfish is described in which reflex activity is maintained for eight hours or more. 2. Intracellular recording and cobalt injection have made it possible to locate and characterize the claw motoneurons. Soma recordings show attenuated axon spikes of 5–10 mV amplitude and subthreshold potentials of the same magnitude. 3. The fast closer excitor (FCE) receives subthreshold excitation in parallel with the slow closer excitor (SCE) and the opener inhibitor (OI) even though it seldom fires. 4. IPSP's are recorded in the opener excitor (OE) concurrent with OI spikes. 5. Cobalt injection reveals a parallel dendritic structure among the three synergists FCE, SCE and OI, and a distinctly different dendritic distribution for OE. The soma siza ranking: OI< OE≦FCE = SCE is apparent. 6. The results are discussed and compared with other arthropod systems with regard to the relation between soma size and electrical functions, distribution of somata and dendrites, and effects of cobalt on electrical functioning. A conflict with previous work on this system is discussed.