Postganglionic sympathetic neurons, but not locus coeruleus optostimulation, activates neuromuscular transmission in the adult mouse in vivo.

Recent work demonstrated that sympathetic neurons innervate the skeletal muscle near the neuromuscular junction (NMJ), and muscle sympathectomy and sympathomimetic agents strongly influence motoneuron synaptic vesicle release ex vivo. Moreover, reports attest that the pontine nucleus locus coeruleus (LC) projects to preganglionic sympathetic neurons and regulates human mobility and skeletal muscle physiology. Thus, we hypothesized that peripheral and central sympathetic neurons projecting directly or indirectly to the skeletal muscle regulate NMJ transmission. The aim of this study was to define the specific neuronal groups in the peripheral and central nervous systems that account for such regulation in adult mice in vivo by using optogenetics and NMJ transmission recordings in 3-5-month-old, male and female ChR2(H134R/EYFP)/TH-Cre mice. After detecting ChR2(H134R)/EYFP fluorescence in the paravertebral ganglia and LC neurons, we tested whether optostimulating the plantar nerve near the lumbricalis muscle or LC neurons effectively modulates motor nerve terminal synaptic vesicle release in living mice. Nerve optostimulation increased motor synaptic vesicle release in vitro and in vivo, while the presynaptic adrenoceptor blockers propranolol (ß1/ß2) and atenolol (ß1) prevented this outcome. The effect is primarily presynaptic since miniature end-plate potential (MEPP) kinetics remained statistically unmodified after stimulation. In contrast, optostimulation of LC neurons did not regulate NMJ transmission. In summary, we conclude that postganglionic sympathetic neurons, but not LC neurons, increased NMJ transmission by acting on presynaptic ß1-adrenergic receptors in vivo.

Magnetic Fields Modulate Blue-Light-Dependent Regulation of Neuronal Firing by Cryptochrome.

Many animals are able to sense the Earth's geomagnetic field to enable behaviors such as migration. It is proposed that the magnitude and direction of the geomagnetic field modulates the activity of cryptochrome (CRY) by influencing photochemical radical pair intermediates within the protein. However, this proposal will remain theoretical until a CRY-dependent effect on a receptor neuron is shown to be modified by an external magnetic field (MF). It is established that blue-light (BL) photoactivation of CRY is sufficient to depolarize and activate Drosophila neurons. Here, we show that this CRY-dependent effect is significantly potentiated in the presence of an applied MF (100 mT). We use electrophysiological recordings from larval identified motoneurons, in which CRY is ectopically expressed, to show that BL-dependent depolarization of membrane potential and increased input resistance are markedly potentiated by an MF. Analysis of membrane excitability shows that these effects of MF exposure evoke increased action potential firing. Almost nothing is known about the mechanism by which a magnetically induced change in CRY activity might produce a behavioral response. We further report that specific structural changes to the protein alter the impact of the MF in ways that are strikingly similar to those from recent behavioral studies into the magnetic sense of Drosophila These observations provide the first direct experimental evidence to support the hypothesis that MF modulation of CRY activity is capable of influencing neuron activity to allow animal magnetoreception. SIGNIFICANCE STATEMENT: The biophysical mechanism of animal magnetoreception is still unclear. The photoreceptor protein cryptochrome has risen to prominence as a candidate magnetoreceptor molecule based on multiple reports derived from behavioral studies. However, the role of cryptochrome as a magnetoreceptor remains controversial primarily because of a lack of direct experimental evidence linking magnetic field (MF) exposure to a change in neuronal activity. Here, we show that exposure to an MF (100 mT) is sufficient to potentiate the ability of light-activated cryptochrome to increase neuronal action potential firing. Our results provide critical missing evidence to show that the activity of cryptochrome is sensitive to an external MF that is capable of modifying animal behavior.

Co-expression of VAL- and TMT-opsins uncovers ancient photosensory interneurons and motorneurons in the vertebrate brain.

The functional principle of the vertebrate brain is often paralleled to a computer: information collected by dedicated devices is processed and integrated by interneuron circuits and leads to output. However, inter- and motorneurons present in today's vertebrate brains are thought to derive from neurons that combined sensory, integration, and motor function. Consistently, sensory inter-motorneurons have been found in the simple nerve nets of cnidarians, animals at the base of the evolutionary lineage. We show that light-sensory motorneurons and light-sensory interneurons are also present in the brains of vertebrates, challenging the paradigm that information processing and output circuitry in the central brain is shielded from direct environmental influences. We investigated two groups of nonvisual photopigments, VAL- and TMT-Opsins, in zebrafish and medaka fish; two teleost species from distinct habitats separated by over 300 million years of evolution. TMT-Opsin subclasses are specifically expressed not only in hypothalamic and thalamic deep brain photoreceptors, but also in interneurons and motorneurons with no known photoreceptive function, such as the typeXIV interneurons of the fish optic tectum. We further show that TMT-Opsins and Encephalopsin render neuronal cells light-sensitive. TMT-Opsins preferentially respond to blue light relative to rhodopsin, with subclass-specific response kinetics. We discovered that tmt-opsins co-express with val-opsins, known green light receptors, in distinct inter- and motorneurons. Finally, we show by electrophysiological recordings on isolated adult tectal slices that interneurons in the position of typeXIV neurons respond to light. Our work supports "sensory-inter-motorneurons" as ancient units for brain evolution. It also reveals that vertebrate inter- and motorneurons are endowed with an evolutionarily ancient, complex light-sensory ability that could be used to detect changes in ambient light spectra, possibly providing the endogenous equivalent to an optogenetic machinery.

Photo-inducible cell ablation in Caenorhabditis elegans using the genetically encoded singlet oxygen generating protein miniSOG.

We describe a method for light-inducible and tissue-selective cell ablation using a genetically encoded photosensitizer, miniSOG (mini singlet oxygen generator). miniSOG is a newly engineered fluorescent protein of 106 amino acids that generates singlet oxygen in quantum yield upon blue-light illumination. We transgenically expressed mitochondrially targeted miniSOG (mito-miniSOG) in Caenorhabditis elegans neurons. Upon blue-light illumination, mito-miniSOG causes rapid and effective death of neurons in a cell-autonomous manner without detectable damages to surrounding tissues. Neuronal death induced by mito-miniSOG appears to be independent of the caspase CED-3, but the clearance of the damaged cells partially depends on the phagocytic receptor CED-1, a homolog of human CD91. We show that neurons can be killed at different developmental stages. We further use this method to investigate the role of the premotor interneurons in regulating the convulsive behavior caused by a gain-of-function mutation in the neuronal acetylcholine receptor acr-2. Our findings support an instructive role for the interneuron AVB in controlling motor neuron activity and reveal an inhibitory effect of the backward premotor interneurons on the forward interneurons. In summary, the simple inducible cell ablation method reported here allows temporal and spatial control and will prove to be a useful tool in studying the function of specific cells within complex cellular contexts.

[Muscular strength development by electromagnetic stimulation].

A new tool for muscular strength development by electromagnetic stimulation (MS) of muscular during voluntary contraction has been described. 18 healthy subjects (men) took part in the research. They were devided into two groups--control (CG) and experimental (EG). Subjects of CG and EG have equal muscular strength parameters. M. gastrocnemius of subjects in EG was exposed to MS (1.8 T, 5 Hz) during training exercises (plantar foot flection). The subjects of CG did not receive MS. The torque of plantar foot flection of EG subjects increased significantly (24%) during 10 days training. The torque of plantar foot flection of CG subjects did not change significantly. We hypothesize increasing of muscular strength of EG subjects was result of high-threshold motor units activation under MS.

A threshold dose of heavy ion radiation that decreases the oxidative enzyme activity of spinal motoneurons in rats.

The effect of heavy ion radiation exposure of the spinal cord on the properties of the motoneurons innervating the slow soleus and fast plantaris muscles was investigated. A 15-, 20-, 40-, 50-, or 70-Gy dose of carbon ions (5 Gy/min) was applied to the 2nd to the 6th lumbar segments of the spinal cord in rats. After a 1-month recovery period, the number and cell body size of the irradiated motoneurons innervating the soleus and plantaris muscles did not differ from that of the non-irradiated controls, irrespective of the dose received. However, the oxidative enzyme activity of these motoneurons was decreased by heavy ion radiation at doses of 40, 50, and 70 Gy compared to that of the non-irradiated controls. This decrease in oxidative enzyme activity levels in the motoneurons returned to that of the non-irradiated controls after a 6-month recovery period. We conclude that heavy ion radiation at doses of 40-70 Gy reversibly decreases the oxidative enzyme activity of motoneurons in the spinal cord of rats.

Mechanism behind the neuronal ephaptic coupling during synchronizing by specific brain-triggered wave as neuronal motor toolkit.

Probable mechanism behind the neuronal ephaptic coupling is investigated based on the introduction of "Brain"-triggered potential excitation signal smartly with a specific very low frequency (VLF) waves as a neuronal motor toolkit. Detection of this electric motor toolkit is attributed to in-vitro precise analyses of a neural network of snail, along to the disconnected snail's neuronal network as a control. This is achieved via rapid (real-time) electrical signals acquisition by blind patch-clamp method during micro-electrode implanting in the neurons at the gigaseal conditions by the surgery operations. This process is based on its waveform (potential excitation signal) detection by mathematical curve fitting process. The characterized waveform of this electrical signal is "Saw Tooth" that is smartly stimulated, alternatively, by the brain during triggering the action potential's (AP's) hyperpolarization zone at a certain time interval at the several µs levels. Triggering the neuron cells results in (1) observing a positive shift (10.0%, depending on the intensity of the triggering wave), and (2) major promotion in the electrical current from sub nano (n) to micro (µ) amper (nA, µA) levels. Direct tracing the time domain (i.e., electrical signal vs. time) and estimation of the frequency domain (diagram of electrical response vs. the applied electrical frequencies) by the "Discrete Fast Fourier Transform" algorithm approve the presence of bilateral and reversible electrical currents between axon and dendrite. This mechanism therefore opens a novel view about the neuronal motor toolkit mechanism, versus the general knowledge about the unilateral electrical current flow from axon to dendrite operations in as neural network. The reliability of this mechanism is evaluated via (1) sequential modulation and demodulation of the snail's neuron network by a simulation electrical functions and sequentially evaluation of the neuronal current sensitivity between pA and nA (during the promotion of the signal-to-noise ratio, via averaging of 30 ± 1 (n = 15) and recycling the electrical cycles before any neuronal response); and (2) operation of the process on the differentiated stem cells. The interstice behavior is attributed to the effective role of Ca2+ channels (besides Na+ and K+ ionic pumping), during hyper/hypo calcium processes, evidenced by inductively coupled plasma as the selected analytical method. This phenomenon is also modeled during proposing quadrupole well potential levels in the neuron systems. This mechanism therefore points to the microprocessor behavior of neuron networks. Stimulation of the neuronal system based on this mechanism, not only controls the sensitivity of neuron electrical stimulation, but also would open a light window for more efficient operating the neuronal connectivity during providing interruptions by phenomena such as neurolysis as well as an efficient treatment of neuron-based disorders.

Thermal laser ablation with tunable lesion size reveals multiple origins of seizure-like convulsions in Caenorhabditis elegans.

Laser microsurgery has long been an important means of assessing the functions of specific cells and tissues. Most laser ablation systems use short, highly focused laser pulses to create plasma-mediated lesions with dimensions on the order of the wavelength of light. While the small size of the lesion enables ablation with high spatial resolution, it also makes it difficult to ablate larger structures. We developed an infrared laser ablation system capable of thermally lesioning tissues with spot sizes tunable by the duration and amplitude of laser pulses. We used our laser system in the roundworm C. elegans to kill single neurons and to sever the dorsal and ventral nerve cords, structures that are difficult to lesion using a plasma-based ablation system. We used these ablations to investigate the source of convulsions in a gain-of-function mutant for the acetylcholine receptor ACR-2. Severing the ventral nerve cord caused convulsions to occur independently anterior and posterior to the lesion, suggesting that convulsions can arise independently from distinct subsets of the motor circuit.

Effect of Pulsed and Continuous Ultrasound Therapy on the Degree of Collateral Axonal Branching at the Lesion Site, Polyinnervation of Motor End Plates, and Recovery of Motor Function after Facial Nerve Reconstruction.

The aim of the present study is to test whether ultrasound therapy of muscles denervated by nerve injury would improve the quality of their reinnervation by reduction of the collateral axonal branching at the lesion site and poly-innervation degree at the neuromuscular junctions. After transection and suture of the buccal branch of the facial nerve, pulsed or continuous type of ultrasound therapy was applied to the paralyzed whisker pad muscles of rats in the course of 2 months. Instead of reduction, we found a significant increase in the collateral axonal branching after continuous ultrasound therapy when compared to the branching determined after pulsed or sham ultrasound therapy. Both types of ultrasound therapy also failed to reduce the proportion of polyinnervated end plates in the reinnervated facial muscles. Accordingly, continuous ultrasound therapy failed to restore any parameter of the motor performance of the vibrissal hairs. Application of pulsed ultrasound therapy promoted slight improvements of the functional parameters angular velocity and acceleration. The inhomogeneous structural and functional results achieved after both types of ultrasound therapy let us conclude that further studies are required to evaluate its effects on peripheral nerve regeneration. Anat Rec, 302:1314-1324, 2019. © 2019 Wiley Periodicals, Inc.

Investigation of the neuroprotective effects of thymoquinone on rat spinal cord exposed to 900 MHz electromagnetic field.

Exposure to electromagnetic field in long-term use of cell phones has increased concerns about serious health problems. Our aim was to survey the possible effects of electromagnetic field radiation (60 min/day for 28 days) on the spinal cords of 12 weeks old rats. Further, we investigated whether the administration of thymoquinone (10 mg/kg/day) would protect the spinal cord tissue against the adverse effects of electromagnetic field or not. Twenty-four adult male Wistar albino rats were assigned randomly into four groups: control, electromagnetic field, thymoquinone and electromagnetic field + thymoquinone. The cervical spinal cords of all rats was evaluated using the stereological, biochemical and histological methods. The number of motor neurons were reduced in the electromagnetic field group compared to the control group (p < 0.05). Superoxide dismutase level was higher in the electromagnetic field group compared to the control group (p < 0.05). In the electromagnetic field + thymoquinone group, we found an increase in the number of motor neurons and decrease in superoxide dismutase activity compared to the electromagnetic field group (p < 0.05). Our histological findings also exhibit the remarkable architectural alterations. We speculated that electromagnetic field radiation induced the morphological and biochemical damage to the spinal cord tissue of rats. Administration of antioxidant, thymoquinone, also ameliorated such complications caused by electromagnetic field.

Segmental, synaptic actions of commissural interneurons in the mouse spinal cord.

Left-right alternation depends on activity in commissural interneurons (CINs) that have axons crossing in the midline. In this study, we investigate the CIN connectivity to local motor neurons using a newly developed preparation of the in vitro neonatal mouse spinal cord that allows us to identify all classes of CINs. Nineteen of 29 short-range CINs with axonal projections <1.5 segments (sCINs) directly excited, directly inhibited, or indirectly inhibited contralateral motor neurons in the quiescent spinal cord. Excitation was glutamatergic and inhibition was mixed glycinergic and/or GABAergic. Long-range CINs were also found to have input to local, contralateral motor neurons. Thirteen of 29 descending CINs had similar synaptic connectivity to contralateral motor neurons as the sCINs, including direct excitation and direct and indirect inhibition. Some (9 of 23) rostrally projecting ascending CINs, and a few (2 of 10) CINs with bifurcating axons that both ascend and descend, indirectly inhibited local, contralateral motor neurons. Rhythmic firing during locomotor-like activity was observed in a number of CINs with segmental synaptic effects on contralateral motor neurons. This study outlines the basic connectivity pattern of CINs in the mouse spinal cord on a segmental level. Our study suggests that, based on observed synaptic connectivity, both short- and long-range CINs are likely involved in segmental left-right coordination and that the CIN system is organized into a dual-inhibitory and single-excitatory system. These systems are organized in a way that they could provide appropriate coordination during locomotion.

DNA topoisomerase I inhibitors ameliorate seizure-like behaviors and paralysis in a Drosophila model of epilepsy.

The Drosophila DNA topoisomerase type I mutant allele, top1JS is an effective general seizure-suppressor mutation, reverting seizure-sensitive phenotypes of several mutant strains in a genetic model of epilepsy. Seizure-suppression is caused by reduced transcription of the top1 (topoisomerase I gene) gene [Song J, Hu J, Tanouye MA. (2007) Seizure suppression by top1 mutations in Drosophila. J Neurosci 27(11):2927-2937]. Here, we examine the possibility that pharmaceutical inhibition of Top1 (topoisomerase I protein) enzymatic activity may also be effective at reducing seizure phenotypes. We investigate the effect of vertebrate Top1 inhibitor camptothecin (CPT) along with two related compounds, apigenin and kaempferol, when fed to seizure-sensitive mutant Drosophila. All three Top1 inhibitors were found to suppress phenotypes in these mutants. In particular, for drug treatments, the recovery time from seizure and paralysis is greatly reduced compared with untreated animals. Intriguingly we find that chronic drug treatments result in a small reduction in seizure sensitivity. Taken together, the results suggest that Top1 inhibitors may have the potential to be developed into effective anti-epileptic drugs, especially for brain tumor patients presenting with epilepsy.

A chronically implantable device for the controlled delivery of substances, and stimulation and recording of activity in severed nerves.

We describe the use of an implantable device for peripheral nerves that allows chronic simultaneous delivery of small volumes of solution, recording of both field and multiunit potentials, and electrical stimulation. This custom-made multifunctional device was attached to the cut end of the abducens (VIth) nerve for stimulation, recording and injection purposes. Our device consists of a polyethylene chamber with two electrodes that can be used for stimulation and recording and two Teflon tubes that serve as inlet and outlet for administering chemicals to the nerve fitted inside. Since the device is implanted in a retro-orbital position, we herein will refer to it as an intraorbitary device (IOD). The applicability of the IOD is demonstrated with an electrophysiological and anatomical account of the properties of the abducens nerve. Furthermore, it is shown that certain neuronal discharge properties can be inferred from the nerve recordings. The IOD can also be efficiently used for the delivery of small volume of pharmacological substances or conventional retrograde markers.

In vivo evidence for serotonin 5-HT2C receptor-mediated long-lasting excitability of lumbar spinal reflex and its functional interaction with 5-HT1A receptor in the mammalian spinal cord.

The purpose of the present study was to investigate the 5-HT(2C) receptor-mediated effects on the spinal monosynaptic mass reflex activities and also its functional interactions with 5-HT(1A) receptors in anesthetized, acutely spinalized mammalian adult spinal cord in vivo. Intravenous administration of (+/-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI) (0.1 mg/kg), an agonist of 5-HT(2A/2C) receptors, significantly increased the excitability of spinal motoneurons as reflected by an increase in the spinal monosynaptic mass reflex amplitude to 150-200% of the control. 5-HT(2A/2C) receptor-induced motoneuron excitability was slow, persistent and long-lasting for more than 2h that was significantly inhibited by 5-HT(2C) receptor specific antagonist SB 242084 administered 10 min prior to DOI. Simultaneous administration of DOI (0.1 mg/kg, i.v.) along with (+/-)-8-hydroxy dipropylaminotetraline hydrobromide (8-OH-DPAT) (0.1 mg/kg, i.v.) completely inhibited DOI-induced spinal monosynaptic mass reflex facilitation. In another separate study, administration of 8-OH-DPAT (0.1 mg/kg, i.v.) at the maximum response of DOI also inhibited the motoneuron's excitability; however, the inhibition lasted only for a period of 40-60 min after administration of 8-OH-DPAT, after which the spinal monosynaptic mass reflex amplitude reached its maximum level. These findings suggest that the 5-HT(2C) receptor is primarily involved in the mediation of the long-lasting excitability of spinal motoneurons and possibly interacts with its functional counterpart, 5-HT(1A) receptors in the mammalian adult spinal cord.

Coordination of eye and head components of movements evoked by stimulation of the paramedian pontine reticular formation.

Constant frequency microstimulation of the paramedian pontine reticular formation (PPRF) in head-restrained monkeys evokes a constant velocity eye movement. Since the PPRF receives significant projections from structures that control coordinated eye-head movements, we asked whether stimulation of the pontine reticular formation in the head-unrestrained animal generates a combined eye-head movement or only an eye movement. Microstimulation of most sites yielded a constant-velocity gaze shift executed as a coordinated eye-head movement, although eye-only movements were evoked from some sites. The eye and head contributions to the stimulation-evoked movements varied across stimulation sites and were drastically different from the lawful relationship observed for visually-guided gaze shifts. These results indicate that the microstimulation activated elements that issued movement commands to the extraocular and, for most sites, neck motoneurons. In addition, the stimulation-evoked changes in gaze were similar in the head-restrained and head-unrestrained conditions despite the assortment of eye and head contributions, suggesting that the vestibulo-ocular reflex (VOR) gain must be near unity during the coordinated eye-head movements evoked by stimulation of the PPRF. These findings contrast the attenuation of VOR gain associated with visually-guided gaze shifts and suggest that the vestibulo-ocular pathway processes volitional and PPRF stimulation-evoked gaze shifts differently.

Fatigue-sensitive afferents inhibit extensor but not flexor motoneurons in humans.

The role of group III and IV muscle afferents in controlling the output from human muscles is poorly understood. We investigated the effects of these afferents from homonymous or antagonist muscles on motoneuron pools innervating extensor and flexor muscles of the elbow. In study 1, subjects (n = 8) performed brief maximal voluntary contractions (MVCs) of elbow extensors before and after a 2 min MVC of the extensors. During MVCs, electromyographic responses from triceps were evoked by stimulation of the corticospinal tracts [cervicomedullary motor evoked potentials (CMEPs)]. The same subjects repeated the protocol, but input from fatigue-sensitive afferents was prolonged after the fatiguing contraction by maintained muscle ischemia. In study 2, CMEPs were evoked in triceps during brief extensor MVCs before and after a 2 min sustained flexor MVC (n = 7) or in biceps during brief flexor MVCs before and after a sustained extensor MVC (n = 7). Again, ischemia was maintained after the sustained contractions. During sustained MVCs of the extensors, CMEPs in triceps decreased by approximately 35%. Without muscle ischemia, CMEPs recovered within 15 s, but with maintained ischemia, they remained depressed (by approximately 28%; p < 0.001). CMEPs in triceps were also depressed (by approximately 20%; p < 0.001) after fatiguing flexor contractions, whereas CMEPs in biceps were facilitated (by approximately 25%; p < 0.001) after fatiguing extensor contractions. During fatigue, inputs from group III and IV muscle afferents from homonymous or antagonist muscles depress extensor motoneurons but facilitate flexor motoneurons. The more pronounced inhibitory influence of these afferents on extensors suggests that these muscles may require greater cortical drive to generate force during fatigue.

Distinct physiological mechanisms underlie altered glycinergic synaptic transmission in the murine mutants spastic, spasmodic, and oscillator.

Spastic (spa), spasmodic (spd), and oscillator (ot) mice have naturally occurring glycine receptor (GlyR) mutations, which manifest as motor deficits and an exaggerated "startle response." Using whole-cell recording in hypoglossal motoneurons, we compared the physiological mechanisms by which each mutation alters GlyR function. Mean glycinergic miniature IPSC (mIPSC) amplitude and frequency were dramatically reduced (>50%) compared with controls for each mutant. mIPSC decay times were unchanged in spa/spa (4.5 +/- 0.3 vs 4.7 +/- 0.2 ms), reduced in spd/spd (2.7 +/- 0.2 vs 4.7 +/- 0.2 ms), and increased in ot/ot (12.3 +/- 1.2 vs 4.8 +/- 0.2 ms). Thus, in spastic, GlyRs are functionally normal but reduced in number, whereas in spasmodic, GlyR kinetics is faster. The oscillator mutation results in complete absence of alpha1-containing GlyRs; however, some non-alpha1-containing GlyRs persist at synapses. Fluctuation analysis of membrane current, induced by glycine application to outside-out patches, showed that mean single-channel conductance was increased in spa/spa (64.2 +/- 4.9 vs 36.1 +/- 1.4 pS), but unchanged in spd/spd (32.4 +/- 2.1 vs 35.3 +/- 2.1 pS). GlyR-mediated whole-cell currents in spa/spa exhibited increased picrotoxin sensitivity (27 vs 71% block for 100 microM), indicating alpha1 homomeric GlyR expression. The picrotoxin sensitivity of evoked glycinergic IPSCs and conductance of synaptic GlyRs, as determined by nonstationary variance analysis, were identical for spa/spa and controls. Together, these findings show the three mutations disrupt GlyR-mediated inhibition via different physiological mechanisms, and the spastic mutation results in "compensatory" alpha1 homomeric GlyRs at extrasynaptic loci.

Changes of motor unit contractile output during repeated activity.

The aim of the study was to evaluate changes in the motor unit output and to determine changes in the optimal stimulation frequency (i.e., giving the maximal output per one pulse) during prolonged contractile activity when, successively, potentiation of force and fatigue developed. The influence of these phenomena was studied on three types of motor units: fast fatigable (FF), fast resistant (FR) and slow (S) in the rat medial gastrocnemius muscle. The motor units were isolated by a method of splitting of L5 ventral root into very thin bundles of axons which were electrically stimulated 10 times with repeated series of 10 trains of stimuli at duration of 500 ms and progressively increasing (1-150 Hz) frequency. The initial (the first series of stimulating trains), potentiated (the second series), as well as fatigued (the tenths series) force recordings were compared. The motor unit output was expressed as the area under the force-time record in response to one stimulus measured at a plateau phase of the tetanic force. The stimulation frequency when the force-time area per one pulse was maximal was accepted as the optimal frequency. In fast motor units, the maximal contractile output increased with potentiation and was reduced with fatigue, and the optimal frequency decreased and increased, respectively. Nevertheless, the fusion degrees of the optimal tetanic contractions were similar in initial state, potentiation and fatigue independently of the changes in force. The applied stimulation protocol had almost no influence on the mechanical activity of slow motor units. The study highlights the physiological importance of force potentiation induced by preceding contractile activity for the economy of motor performance. The observed changes of the optimal stimulation frequency are consistent with the known changes in the motor unit firing rates during voluntary activity when the two phenomena develop.

Effects of repetitive transcranial magnetic stimulation on masseter motor-neuron pool excitability.

OBJECTIVE: Repetitive transcranial magnetic stimulation (rTMS) has been widely used to modulate the excitability of the cortical control of limbs muscles, but rarely in the cortical control of human masseter muscles. This study aims to investigate the effects of rTMS on masseter motor-neuron pool excitability in humans. MATERIALS AND METHODS: A total of 20 healthy participants were selected and received a total of three rTMS sessions involving stimulation of the right masseter-motor complex: one session of 10-Hz rTMS, one session of 1-Hz rTMS and one session of sham rTMS at an intensity of 80% of the active motor threshold (AMT). The masseter AMT, motor-evoked potentials (MEPs), cortical-silent period (CSP), and short-interval intracortical inhibition (SICI) were measured before and after each rTMS session. RESULTS: The masseter SICI was significantly decreased following 10-Hz rTMS, with no significant changes in AMT, MEPs or CSP. No significant differences in masseter AMT, MEPs, CSP or SICI were observed in either the 1-Hz, or sham rTMS groups. CONCLUSIONS: The present findings demonstrate that high-frequency rTMS increases masseter motor-neuron pool excitability.

Regulation of motoneuron excitability via motor endplate acetylcholine receptor activation.

Motoneuron populations possess a range of intrinsic excitability that plays an important role in establishing how motor units are recruited. The fact that this range collapses after axotomy and does not recover completely until after reinnervation occurs suggests that muscle innervation is needed to maintain or regulate adult motoneuron excitability, but the nature and identity of underlying mechanisms remain poorly understood. Here, we report the results of experiments in which we studied the effects on rat motoneuron excitability produced by manipulations of neuromuscular transmission and compared these with the effects of peripheral nerve axotomy. Inhibition of acetylcholine release from motor terminals for 5-6 d with botulinum toxin produced relatively minor changes in motoneuron excitability compared with the effect of axotomy. In contrast, the blockade of acetylcholine receptors with alpha-bungarotoxin over the same time interval produced changes in motoneuron excitability that were statistically equivalent to axotomy. Muscle fiber recordings showed that low levels of acetylcholine release persisted at motor terminals after botulinum toxin, but endplate currents were completely blocked for at least several hours after daily intramuscular injections of alpha-bungarotoxin. We conclude that the complete but transient blockade of endplate currents underlies the robust axotomy-like effects of alpha-bungarotoxin on motoneuron excitability, and the low level of acetylcholine release that remains after injections of botulinum toxin inhibits axotomy-like changes in motoneurons. The results suggest the existence of a retrograde signaling mechanism located at the motor endplate that enables expression of adult motoneuron excitability and depends on acetylcholine receptor activation for its normal operation.