Excitatory neural control of posterograde heartbeat by the frontal ganglion in the last instar larva of a lepidopteran, Bombyx mori

The frontal ganglion of the silkworm (Bombyx mori) gives rise to a visceral nerve, branches of which include a pair of anterior cardiac nerves and a pair of the posterior cardiac nerves. Forward-fill of the visceral nerve with dextran labeled with tetramethyl rhodamine shows the anterior cardiac nerves innervate the anterior region of the dorsal vessel. Back-fill of the anterior cardiac nerves with Co²⁺ and Ni²⁺ ions and the fluorescent dye reveals that the cell bodies of two motor neurons are located in the frontal ganglion. Injection of 5, 6-carboxyfluorescein into the cell body of an identified motor neuron shows that the neuron gives rise to an axon running to the visceral nerve. Unitary excitatory junctional potentials (EJPs) were recorded from a myocardial cell at the anterior end of the heart. They responded in a one-to-one manner to electrical stimuli applied to the visceral nerve, or to impulses generated by a depolarizing current injected into the cell body. EJPs induced by stimuli at higher than 0.5 Hz showed facilitation while those induced at higher than 2 Hz showed summation. Individual EJPs without summation, or a train of EJPs with summation, caused acceleration in the phase of posterograde heartbeat and heart reversal from anterograde heartbeat to posterograde heartbeat. It is likely that the innervation of the anterior region of the dorsal vessel by the motor neurons, through the anterior cardiac nerves is responsible for the control of heartbeat in Lepidoptera, at least in part.     

Transplantation of Xenopus laevis Tissues to Determine the Ability of Motor Neurons to Acquire a Novel Target

The evolutionary origin of novelties is a central problem in biology. At a cellular level this requires, for example, molecularly resolving how brainstem motor neurons change their innervation target from muscle fibers (branchial motor neurons) to neural crest-derived ganglia (visceral motor neurons) or ear-derived hair cells (inner ear and lateral line efferent neurons). Transplantation of various tissues into the path of motor neuron axons could determine the ability of any motor neuron to innervate a novel target. Several tissues that receive direct, indirect, or no motor innervation were transplanted into the path of different motor neuron populations in Xenopus laevis embryos. Ears, somites, hearts, and lungs were transplanted to the orbit, replacing the eye. Jaw and eye muscle were transplanted to the trunk, replacing a somite. Applications of lipophilic dyes and immunohistochemistry to reveal motor neuron axon terminals were used. The ear, but not somite-derived muscle, heart, or liver, received motor neuron axons via the oculomotor or trochlear nerves. Somite-derived muscle tissue was innervated, likely by the hypoglossal nerve, when replacing the ear. In contrast to our previous report on ear innervation by spinal motor neurons, none of the tissues (eye or jaw muscle) was innervated when transplanted to the trunk. Taken together, these results suggest that there is some plasticity inherent to motor innervation, but not every motor neuron can become an efferent to any target that normally receives motor input. The only tissue among our samples that can be innervated by all motor neurons tested is the ear. We suggest some possible, testable molecular suggestions for this apparent uniqueness.     

An electron microscopic study on the innervation of the gill filaments of a lamprey,Lampetra japonica

This paper reports observations on the innervation of gill filaments of the lamprey, Lampetra japonica. Nerve fibers run on each side of the afferent filament artery (AFA nerve) and in the connective tissue compartment along the efferent filament artery (EFA nerve). The AFA nerve supplies vasomotor fibers to the afferent filament artery and arteriovenous anastomoses and special visceral motor fibers to branchial muscle fibers (musculus compressor branchialis circularis). Nerve endings of the vasomotor fibers contain large, cored vesicles (60–180 nm in diameter) with a variable number of small, clear vesicles (30–70 μm in diameter), whereas those of the visceral motor fibers have many small, clear vesicles with few large, cored vesicles. The EFA nerve supplies vasomotor fibers to the efferent filament artery. Their endings, containing mixtures of predominantly large, cored vesicles and small, clear vesicles make close synaptic contacts with reticular cells. The latter in turn are connected with each other or with smooth muscle cells in the wall of the efferent filament artery by nexuses. No nerves are found in the axial plate between the afferent and efferent filament arteries nor in the secondary lamellae of individual gill filaments. No afferent nerve supply to the gill filament has been found.     

Proliferation patterns of dorsal root ganglion neurons of cutaneous,muscle and visceral nerves in the rat

In a previous study we provided evidence that dorsal root ganglion (DRG) neurons of different phenotypes have different birthdates. The present study aimed at determining if birthdates of DRG neurons are related to different types of peripheral nerves, namely cutaneous versus muscle, and somatic versus visceral. Pregnant rats were injected intraperitoneally with bromodeoxyuridine (BrdU) to label the neurons on one of the embryonic days E12–E16. When the progeny rats reached adulthood, a mixture of 1% B-fragment of cholera toxin and 1% isolectin B4 from Griffonia simplicifolia I was injected into the peripheral nerves, or a 5% Fluoro-Gold solution was applied to the transected end of the nerves. The saphenous and sural nerves were used as cutaneous nerves, the gastrocnemius nerve as a muscle nerve, the intercostal nerves T9–11 as somatic nerves and the greater splanchnic nerve as a visceral nerve. Cell size measurements were made of DRG neurons labeled from the two cutaneous nerves and the muscle nerve, as well as of neurons of the saphenous and gastrocnemius nerves labeled by BrdU at different embryonic stages. Most of the DRG neurons of the muscle and intercostal nerves were generated early, with peaks at E13, and those of the cutaneous and visceral afferent nerves later, with peaks at E14. The temporal differences were reflected in the cell size spectrum, the muscle nerve having a greater proportion of large neurons compared to the cutaneous nerves. The findings add to previous knowledge regarding the sequence of development of different DRG phenotypes.     

EFFECTS OF BOTULINUM AND TETANUS TOXINS ON CHOLINE ACETYLTRANSFERASE ACTIVITY IN SKELETAL MUSCLE IN THE MOUSE

Abstract— The effects of botulinum and tetanus toxins on the activity of choline acetyltransferase present in the motor nerve terminals of fast and slow skeletal muscle in the mouse were investigated. There was no change in the activities of choline acetyltransferase in either muscle after the injection of botulinum toxin but tetanus toxin caused a rise in the activity of the enzyme in fast muscle. Botulinum toxin is known to inhibit the release of acetylcholine and whilst neuromuscular transmission is blocked the motor nerves sprout and form new end-plates. Tetanus toxin has been shown to cause hyperactivity of motor neurons. The nerve growth caused by the botulinum toxin did not result in increased choline acetyltransferase levels in the muscles, whereas the synaptic hyperactivity caused by tetanus was associated with increased enzyme levels.     

TRANSMITTER METABOLIZING ENZYMES AND FREE AMINO ACID LEVELS IN SENSORY AND MOTOR NERVES AND GANGLIA OF THE SHORE CRAB (CARCINUS MAENAS)

—The distribution of ChAT (choline acetyltransferase), GAD (glutamate decarboxylase) and acetylcholinesterase in some sensory and motor nerves of the shore crab, Carcinus maenas, has been investigated using micro-assay techniques. ChAT was concentrated in the afferent nerve fibres of the thoracic-coxal muscle receptor as well as in the coxo-basal chordotonal receptor nerve and other leg sensory fibres. GAD was found in leg motor nerves including the promotor and remotor muscle nerves, being undetectable in the sensory nerves. Acetylcholinesterase was found in similar levels in both sensory and motor nerves assayed. Amino acid analysis using a micro-dansylation technique showed that sensory nerves had low GABA levels, whereas the leg nerve including motor fibres had substantially higher GABA concentrations. GAD and GABA were also found in low amounts in the leg promoter mucle, which is consistent with GABA being a neuromuscular transmitter.     

THE LOCUS OF THE ACTION OF VERATRIN

1. In Mnemiopsis veratrin shows two stages of veratrin poisoning. First, inhibition of the beats of the plates which disappears on cutting them away either singly or in small groups. Second, after half an hour mechanical stimulation excites the beat of the plates in the intact veratrinized animal. It is concluded that veratrin acts on nervous tissue and not on the substance of the swimming plates. 2. In Lumbricus, veratrin acts on the ventral nerve cord alone, and not on the muscles and peripheral nerves. 3. In Musca, veratrin first causes opisthotonos, then spasms and extreme flexion of the legs. Decapitation causes these effects to disappear hence veratrin acts on the cerebral ganglia of the fly. 4. Veratrin applied to the sciatic nerve of the frog causes, after a latent period of 20 minutes, irregular contractions of the gastrocnemius which persist for an hour or more. Veratrin is thus a neurophil alkaloid of the first class as well as second and in this way resembles tetraethyl ammonium chloride. 5. If the end of a sciatic nerve is dipped into veratrin solution, then direct stimulation of the gastrocnemius muscle results in contraction with delayed relaxation, although the muscle itself is not subject to the action of veratrin. 6. By means of preparations of the sartorius muscle of the frog it is shown that veratrin acts not on the muscle cells directly but on the nerve fibers. Hence veratrin produces the characteristic muscle curve showing delayed relaxation by its action on the nervous elements.     

Nerve Cross-Bridging to Enhance Nerve Regeneration in a Rat Model of Delayed Nerve Repair

There are currently no available options to promote nerve regeneration through chronically denervated distal nerve stumps. Here we used a rat model of delayed nerve repair asking of prior insertion of side-to-side cross-bridges between a donor tibial (TIB) nerve and a recipient denervated common peroneal (CP) nerve stump ameliorates poor nerve regeneration. First, numbers of retrogradely-labelled TIB neurons that grew axons into the nerve stump within three months, increased with the size of the perineurial windows opened in the TIB and CP nerves. Equal numbers of donor TIB axons regenerated into CP stumps either side of the cross-bridges, not being affected by target neurotrophic effects, or by removing the perineurium to insert 5-9 cross-bridges. Second, CP nerve stumps were coapted three months after inserting 0-9 cross-bridges and the number of 1) CP neurons that regenerated their axons within three months or 2) CP motor nerves that reinnervated the extensor digitorum longus (EDL) muscle within five months was determined by counting and motor unit number estimation (MUNE), respectively. We found that three but not more cross-bridges promoted the regeneration of axons and reinnervation of EDL muscle by all the CP motoneurons as compared to only 33% regenerating their axons when no cross-bridges were inserted. The same 3-fold increase in sensory nerve regeneration was found. In conclusion, side-to-side cross-bridges ameliorate poor regeneration after delayed nerve repair possibly by sustaining the growth-permissive state of denervated nerve stumps. Such autografts may be used in human repair surgery to improve outcomes after unavoidable delays.     

Nonacceptance of innervation by innervated neonatal rat muscle

Denervated adult muscle accepts innervation and has high levels of extrajunctional acetylcholine (ACh) receptor, compared to innervated adult muscle. If the high receptor density or any externally oriented part of the receptor molecule permitted or triggered functional synaptogenesis, then innervated neonatal muscle, with its known high extrajunctional sensitivity, should also accept extra synapses from implanted motor nerves. This prediction was tested by implanting the common peroneal nerve into innervated lateral gastrocnemius muscle in 25 neonatal rats and studying the innervation achieved 1–8 weeks later. With one exception, zero or negligible twitch tensions were obtained when the implanted nerve was stimulated. Intracellular recording in two cases showed no evidence of subthresholdevoked potentials in surface muscle fibers. In contrast, when the original motor nerve was cut at the time of common peroneal nerve implantation, reinnervation occurred as soon as 4 days later, and substantial indirect twitches (most observed qualitatively) were invariably found 6–7 days after operation. Four to eight weeks after nerve implantation into denervated muscle, substantial twitch tensions were obtained upon stimulation of the implanted nerve. α-Bungarotoxin binding to extrajunctional ACh receptors per unit surface area was similar in innervated neonatal and denervated adult muscle. Therefore, nonacceptance of additional functional innervation in neonatal muscle implies that a high average density of extrajunctional ACh receptor is not sufficient to permit or trigger functional neuromuscular junction formation.     

Role of the visceral nerve in heart rate variations during different behavioral patterns in Megalobulimus sanctipauli (mollusca,gastropoda, pulmonata)

• 1.1. The role of the visceral nerve in mediating the changes in heart rate associated with different behavioral patterns was investigated in Megalobulimus sanctipauli.
• 2.2. The results of acute and chronic denervation experiments indicate that the visceral nerve has no excitatory or inhibitory tonic action on the heart of snails retracted into the shell, nor does it account for the increase in heart rate associated with the locomotion and feeding behaviors.
• 3.3. These changes in heart rate are, probably, indirect effects of increased activity such as an increase in venous return.
• 4.4. The visceral nerve is responsible for approximately 3/4 of the increase in heart rate associated with the first minute of extrusion.
• 5.5. The small increase in heart rate observed in denervated animals is probably caused by an increase in venous return generated by muscle activity that forces the head and food out of the shell.

     

Long-term changes in neurotrophic factor expression in distal nerve stump following denervation and reinnervation with motor or sensory nerve

Several factors have been proposed to account for poor motor recovery after prolonged denervation, including motor neuron cell death and incomplete or poor regeneration of motor fibers into the muscle. Both may result from failure of the muscle and the distal motor nerve stump to continue expression of neurotrophic factors following delayed muscle reinnervation. This study investigated whether regenerating motor or sensory axons modulate distal nerve neurotrophic factor expression. We found that transected distal tibial nerve up-regulated brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) mRNA, down-regulated neurotrophin-3 and ciliary neurotrophic factor mRNA, and that although these levels returned to normal with regeneration, the chronically denervated distal nerve stump continued to express these neurotrophic factors for at least 6 months following injury. A sensory nerve (the cutaneous saphenous nerve) sutured to distal tibial nerve lowered injury-induced BDNF and GDNF mRNA levels in distal stump, but repair with a mixed nerve (peroneal, containing muscle and cutaneous axons) was more effective. Repair with sensory or mixed nerves did not affect nerve growth factor or neurotrophin-3 expression. Thus, distal nerve contributed to a neurotrophic environment for nerve regeneration for at least 6 months, and sensory nerve repair helped normalize distal nerve neurotrophic factor mRNA expression following denervation. Furthermore, as BDNF and GDNF levels in distal stump increased following denervation and returned to control levels following reinnervation, their levels serve as markers for the status of regeneration by either motor or sensory nerve.     

Age-related characteristics of the myeloarchitectonics of the peripheral nerves

By means of macromicroscopy and microscopy myeloarchitectonics of morphologically and functionally different somatic and visceral nerves (branches: mandibular nerve, cervical spinal nerves, inferior laryngeal nerve, hepatic plexus) have been studied in 11 age groups. During the prenatal period of ontogenesis asynchronism of myelogenesis is stated in various muscle branches of the nerves, dependent on formation of function in corresponding muscles and muscle groups. As demonstrate investigations on peculiarities of myelogenesis course in the somatic and visceral nerves studied, during the period of pre- and postnatal ontogenesis, its dynamics embraces three stages of myelogenesis, determined by G. B. Stovichek for visceral nerves: productive myelogenesis, stages of stabilization and involution. The stage of productive myelogenesis in the somatic nerves studied lasts up to the end of the adolescent period. Two phases are determined in it: the first lasts up to 2-3 years; the second--up to the end of the adolescent period and is characterized with a complete formation of the myelin fibers spectrum. In the visceral nerves studied increase of general amount of myelin fibers and their differentiation are completed simultaneously during the adolescent age. The stabilization stage of myeloarchitectonics of the nerves studied corresponds to the mature age (I and II periods) and the involution stage--to the elderly (and old) age.     

CELLULAR DISTRIBUTION OF 16S ACETYLCHOLINESTERASE

Multiple molecular forms of acetylcholinesterase (AChE; EC 3.1.1.7), in crude extracts of various tissues from the rat, were distinguished by velocity sedimentation analysis on linear sucrose gradients. Skeletal muscle samples containing end-plate regions showed three different forms of AChE with apparent sedimentation coefficients of 16, 10 and 4s. The 16s form was not detected in non-innervated regions of skeletal muscle, large intestine smooth muscle, whole brain tissue, red blood cells or plasma. Spinal cord, a predominantly motor cranial nerve and mixed (sensory and motor) peripheral nerves contained 16, 10, 6.5 and 4S AChE. Ventral motor roots, supplying the sciatic nerve, contained these four forms of the enzyme, while corresponding dorsal sensory roots were devoid of the 16S form. The 16s-AChE confined to ventral roots can be attributed totally to motor neurons and not to Schwann cells composing these roots. Whether the 16s-AChE presently found in motor nerves has chemical identity with that found at motor end-plates is the basis of future experiments.     

The central nervous control of the adductor behaviour of lamellibranch molluscs.

In Egeria radiata (L.) and Mutela dubia (Gmelin) decerebration does not result in tonus of the posterior adductor muscle and each species continues to exhibit its characteristic rapid and slow rhythms. Excitatory and inhibitory nerve pathways originating in the cerebral ganglia terminate on the visceral ganglia. The cerebral ganglia alone do not exhibit any rhythm; the anterior adductor muscle remains relaxed after excision of the visceral ganglia. The mid-dorsal and the anterior lobes of the visceral ganglia in Scrobicularia plana (Da Costa) control all adductor activity. Groups of potentials in the posterior adductor nerves originate from the different lobes and are separate physiological mechanisms.     

BMP4 Is a Peripherally-Derived Factor for Motor Neurons and Attenuates Glutamate-Induced Excitotoxicity In Vitro

Bone morphogenetic proteins (BMPs), members of the transforming growth factor-beta (TGF-β) superfamily, have been shown to play important roles in the nervous system, including neuronal survival and synaptogenesis. However, the physiological functions of BMP signaling in the mammalian neuromuscular system are not well understood. In this study, we found that proteins of the type II bone morphogenetic receptors (BMPRII) were detected at the neuromuscular junction (NMJ), and one of its ligands, BMP4, was expressed by Schwann cells and skeletal muscle fibers. In double-ligated nerves, BMP4 proteins accumulated at the proximal and distal portions of the axons, suggesting that Schwann cell- and muscle fiber-derived BMP4 proteins were anterogradely and retrogradely transported by motor neurons. Furthermore, BMP4 mRNA was down-regulated in nerves but up-regulated in skeletal muscles following nerve ligation. The motor neuron-muscle interactions were also demonstrated using differentiated C2C12 muscle cells and NG108-15 neurons in vitro. BMP4 mRNA and immunoreactivity were significantly up-regulated in differentiated C2C12 muscle cells when the motor neuron-derived factor, agrin, was present in the culture. Peripherally-derived BMP4, on the other hand, promotes embryonic motor neuron survival and protects NG108-15 neurons from glutamate-induced excitotoxicity. Together, these data suggest that BMP4 is a peripherally-derived factor that may regulate the survival of motor neurons.     

Intense ultraterminal sprouting from motor nerves and ultrastructural aspects of the neuromuscular junction and non-junctional sarcolemma of the soleus (slow-twitch) muscle in motor endplate disease in the mouse

Motor end-plate disease (med) in the mouse is an hereditary defect of the neuromuscular system, with partial functional denervation and muscle inactivity in late stages of the disease. Motor end-plate disease is characterized by an intense ultraterminal sprouting of the motor nerves from swollen nerve terminal branches in the soleus muscle. At the ultrastructural level, the neuromuscular junctions extend to very wide territories, often outside the original motor end-plate, in regions where the nerve sprouts are in simple apposition to the muscle fiber, with no secondary synaptic folds. The nerve terminals are rich in neurofilaments and poor in synaptic vesicles.Freeze fracture analysis of the pre-synaptic and post-synaptic membrane specializations fails to reveal any important structural alteration which could suggest a defect in acetylcholine release or in muscle membrane excitability. However, the non-junctional sarcolemmal specializations (the so-called ‘square arrays’) arc found with a frequency slightly higher than in normal muscle.The nerve abnormalities at the neuromuscular junction may be either a consequence of muscle inactivity or the morphological expression of some primary nerve abnormality. Further studies of the soleus muscle at early stages of the disease may provide evidence in favor of either possibility.     

Biogenic amines evoke heartbeat reversal in larvae of the sweet potato hornworm,Agrius convolvuli

The sweet potato hornworm, Agrius convolvuli, possesses a pair of anterior cardiac nerves innervating the dorsal vessel. The anterior cardiac nerves branch off the visceral nerve that arises posteriorly from the frontal ganglion. Heartbeat reversal from anterograde heartbeat to posterograde heartbeat is triggered by the anterior cardiac nerves. Application of octopamine (OA) during the anterograde heartbeat phase reverses the anterograde heartbeat to the posterograde heartbeat, while application of OA during the phase of posterograde heartbeat accelerates heartbeat. The heartbeat reversal from anterograde heartbeat to posterograde heartbeat evoked by stimuli applied to the visceral nerve is blocked by application of the octopaminergic antagonists, phentolamine and chlorpromazine. The results suggest that OA may be a neurotransmitter for the anterior cardiac nerve. The alary muscle of the second segment receives excitatory innervation from the posterior cardiac nerve and from the nerve which extends from the second abdominal ganglion. Activation of the alary muscle results in acceleration of posterograde heartbeat. Other neurotransmitters, besides OA, may take part in the resultant acceleration.     

THE ACTION OF INHIBITORY NERVES ON CARBON DIOXIDE PRODUCTION IN THE HEART GANGLION OF LIMULUS

It has been shown in this paper that stimulation of the inhibitory nerves of the neurogenic heart of Limulus, which correspond to the vagus nerves of the vertebrate heart, results in a marked diminution of CO₂ production in the heart ganglion, while stimulation of the ganglion, leading to increased activity of the heart, leads also to increased CO₂ production by the ganglion. This shows that inhibition of the automaticity of this ganglion by the action of its inhibitory nerves consists, not in a process of blocking, but in a diminution of those chemical reactions in the ganglion cells which give rise to the production of CO₂.     

Influence of nerve supply on hand electromyography coherence during a three-digit task

Intermuscular coupling has been investigated to understand neural inputs to coordinate muscles in a motor performance. However, little is known on the role of nerve innervation on intermuscular coupling. The purpose of this study was to investigate how the anatomy of nerve distribution affected intermuscular coupling in the hand during static grip. Electromyographic (EMG) signals were recorded from intrinsic and extrinsic muscles while subjects performed a static grip. Coherence was computed for muscle pairs innervated by either the same or different nerves. The results did not support the hypothesis that muscles sharing the same nerve exhibit greater coupling than muscles innervated by different nerves. In general, extrinsic muscle pairs displayed higher coherence than intrinsic pairs. The results suggest that intermuscular coupling in a voluntary motor task is likely modulated in a functional manner and that different nerves might transport common neural inputs to functionally coupled muscles.