Smartphone-Enabled Web-Based Simulation of Cellular Neurophysiology for Laboratory Course and its Effectiveness.

The introduction of computer simulations has enhanced the teaching of neurobiology. Many simulators for personal computers are available, but in countries where schools have low school information and communication technology readiness, it is difficult to introduce computer simulations. Even in such countries, however, students often have their own smartphones and are good at operating them. Therefore, we have developed five web-based simulators that cover a wide range of neurophysiology, including single and whole-cell channel currents, membrane potentials and generation and conduction of action potentials using HTML5 and JavaScript. These simulators may be run free of charge on any device, regardless of the model or OS, thereby enabling schools that have no experience in introducing simulations to introduce them easily. These simulators were especially useful in many schools during COVID-19 restrictions. In this paper, we explain the functions of the simulators we have developed and introduce some practical examples. To verify the usefulness of the simulators, we also conducted a survey in the classrooms in which the simulators were used. Understanding and motivation to learn was shown to increase significantly, indicating that these are useful for neurobiology education.

Association of upper and lower airway eosinophilic inflammation with response to omalizumab in patients with severe asthma.

Background: The anti-immunoglobulin E monoclonal antibody, omalizumab, is used to treat severe asthma and has the potential to ameliorate airway inflammation. However, the effect of omalizumab in ameliorating upper airway inflammation has not been fully elucidated. Objective: We investigated the association of upper and lower airway inflammation with the response to omalizumab treatment. Methods: We used the Global Evaluation of Treatment Effectiveness to assess the efficacy of omalizumab in treating 16 patients with severe asthma. We also investigated the symptom score, short-acting ß-agonist inhaler use, pulmonary function, biomarkers, computed tomography scans, and nasal mucosa pathology at omalizumab initiation and after four months of treatment. Results: When the fraction of exhaled nitric oxide (FeNO) and the percentage of sputum eosinophil were used as indicators of lower airway inflammation, positive correlations were found between CD20 B-cell, mast cell, and eosinophil counts in the nasal mucosa. Improved asthma symptoms were observed in 12 of the 16 severe asthma cases. The FeNO and eosinophil levels in the nasal tissue, prior to the administration of omalizumab were predictors of the response to asthma treatment. Conclusions: These findings suggest heterogeneity among people with severe asthma. In addition, the phenotype associated with response to omalizumab, leading to improvement in asthma symptoms, comprises upper airway eosinophilia and high FeNO levels.

Motility of the Labial Palps in Feeding Behavior and its Innervation in the Marine Mussel, Mytilus galloprovincialis.

The labial palps of bivalves are thought to be involved in suspension feeding. However, the function of their muscular movements and neural regulation are still unclear. In semi-intact preparations of Mytilus, in which one valve was removed, suspended particles were removed from the labial palps following two kinds of compound movements: torsional and rotational. Both of these compound movements are therefore thought to function in rejection during feeding. These movements were observed in reduced preparations of isolated labial palps with intact cerebral ganglia, and were maintained even after removal of the cerebral ganglia, suggesting that they are generated by the peripheral neural network. Stimulation of the anterior pallial nerve elicited tetanic contraction of the labial palp, followed by secondary responses, including torsional movement. Secondary responses were dramatically reduced by a high concentration of divalent cations, in which polysynaptic pathways were inhibited. Hence, the cerebral ganglia may play an excitatory role within the peripheral neural network and the labial palp musculature via the anterior pallial nerve. Administration of serotonin induced repetitive muscular movements, whereas dopamine did not induce muscular movements. Serotonin-induced muscular movements were not elicited under a high concentration of divalent cation condition. In histochemical experiments, both the serotonergic and dopaminergic neural processes and cell body-like structures were widely observed inside the labial palp, the anterior pallial nerve, and the cerebral ganglia. Serotonin may thus contribute to activation of polysynaptic peripheral pathways, which are involved in regulating compound movements.

Dopamine D1 Receptor-Mediated Transmission Maintains Information Flow Through the Cortico-Striato-Entopeduncular Direct Pathway to Release Movements.

In the basal ganglia (BG), dopamine plays a pivotal role in motor control, and dopamine deficiency results in severe motor dysfunctions as seen in Parkinson's disease. According to the well-accepted model of the BG, dopamine activates striatal direct pathway neurons that directly project to the output nuclei of the BG through D1 receptors (D1Rs), whereas dopamine inhibits striatal indirect pathway neurons that project to the external pallidum (GPe) through D2 receptors. To clarify the exact role of dopaminergic transmission via D1Rs in vivo, we developed novel D1R knockdown mice in which D1Rs can be conditionally and reversibly regulated. Suppression of D1R expression by doxycycline treatment decreased spontaneous motor activity and impaired motor ability in the mice. Neuronal activity in the entopeduncular nucleus (EPN), one of the output nuclei of the rodent BG, was recorded in awake conditions to examine the mechanism of motor deficits. Cortically evoked inhibition in the EPN mediated by the cortico-striato-EPN direct pathway was mostly lost during suppression of D1R expression, whereas spontaneous firing rates and patterns remained unchanged. On the other hand, GPe activity changed little. These results suggest that D1R-mediated dopaminergic transmission maintains the information flow through the direct pathway to appropriately release motor actions.

Neural pathways to cardioaccelerator neurons in the isopod crustacean Bathynomus doederleini: Cholinergic activation by somatic movements.

We investigated the excitatory and inhibitory input to cardioaccelerator (CA) and cardioinhibitor (CI) neurons located in the thoracic ganglia of the isopod crustacean Bathynomus doederleini by extracellular and intracellular recording. Electrical stimuli applied to the anterior and posterior connectives of single-ganglion preparations, containing either the 2nd or 3rd thoracic ganglion alone, and each of three paired ganglionic nerve roots produced excitatory postsynaptic potentials (EPSPs) in the cell body of a CA neuron. Artificial movements of appendages, such as the thoracic limbs and the swimmerets, also evoked EPSPs in the CA neuron. Electrical stimuli applied to the peripheral nerves running to appendages induced inhibitory postsynaptic potentials (IPSPs) in a CI neuron. Since artificial movements of the appendages caused decrease of CI impulse rate, these IPSPs in the CI neuron may be caused by mechanoproprioceptors in the appendages. Since tachycardia was accompanied by excitation of CA neurons and inhibition of CI neurons, activation of the mechanoproprioceptors may be responsible for tachycardia. EPSPs in CA neurons produced by stimulation of peripheral nerves were augumented by eserinization and blocked by curarization. The activation of CA neurons by ganglionic roots may be mediated by cholinergic processes ascending from mechanoproprioceptors.

The role of the peripheral enteric nervous system in the control of gut motility in the snail Lymnaea stagnalis.

In this study, we asked whether neurons of the enteric nervous system (ENS) isolated from the central nervous system (CNS) are competent to mediate ordered autonomous gut motility (i.e. descending peristalsis, which may be functional for food propulsion down the gut) in the snail Lymnaea stagnalis. Firstly, we explored the origin of autonomous gut contraction in the esophagus and crop. Using extracellular recording, we were able to detect excitatory junctional potentials (EJPs) elicited by motor neuron activity in autonomous rhythmic bursts, showing that neurogenic autonomous contractions existed. We also determined the motor neuron as cholinergic from the antagonistic effects of d-tubocurarine (d-TC) on the EJPs. Interestingly, the "pacemaker region", which drives the rhythm of the automaticity, was found in the crop, which is located distally to the esophagus. Thus, the burst first occurs in the crop and propagates in an ascending direction (i.e. in the opposite direction of the peristalsis) along the esophagus. From the observation of the relationship between the motor neuron activity and the driven motility, proximally decreasing time lag between the neuronal burst and the peak contraction (T-p; Time to peak) was found to be crucial in producing peristalsis. Regional T-p difference was also observed in the electrically evoked contractions in the isolated esophagus. Blocking the motor neuron activity by d-TC attenuated the regional difference. The above suggest that the processes leading up to the elicitation of EJPs by ENS activity contribute to producing the regional T-p difference.

A neural analysis of avoidance conditioning with the feeding attractant glycine in Pleurobranchaea japonica.

Glycine (Gly) is one of the amino acids that most strongly provoke feeding behavior in the carnivorous opisthobranch sea slug Pleurobranchaea japonica. Placing of an aliquot of a Gly solution in front of the anterior end of this animal induced feeding responses such as orientation to the origin of the stimulus and extrusion of the proboscis. In contrast, light stimulation of the body of the animal with a glass-fiber light guide evoked aversive responses involving the gill withdrawal response. Animals were trained by pairing a Gly solution stimulus (conditioned) and a light stimulus (unconditioned). After training with repetitive paired stimuli, we found that animals exhibited aversive responses to the Gly solution. We confirmed achievement of a conditioned Gly-aversive reflex in intact animals by recording of motor impulses with an electrode implanted on the branchial nerve responsible for the gill withdrawal response, the most reliable index of the reflex. Motor discharge of the branchial nerve associated with the conditioned Gly-aversive reflex persisted even in a preparation isolated from an animal which had previously acquired the reflex. This study provides an experimental model for neural analysis with in vivo long-term nerve recording using an electrode implanted in a nerve of an intact animal for neuroethological training.

Verification of the effect of the axon fluid as a highly dielectric medium in the high-speed conduction of action potentials using a novel axon equivalent circuit.

Both sensory neurons and motor neurons transfer signals rapidly through long pathways. Such signals propagate as action potentials through neurons. In myelinated neurons, high conduction velocities of 120 m/s have been reported, even for axons of just 20 µm in diameter. Such a high conduction velocity is enabled by the characteristic morphology of a myelinated axon: repeated regions encased by long uniform myelin sheaths alternating with extremely short exposed regions of the axon called nodes of Ranvier, which generate extremely sharp action potentials. Although the need for the action potential to cross many nodes increases the relay time, it is still able to propagate rapidly. This phenomenon motivated us to derive a new mechanism of the action potential propagation. First, the dielectric effect of the axonal fluid was considered, and it was investigated whether the combination of the characteristic axonal morphology and the dielectric constant of the axonal fluid contributes significantly to the realization of high conduction velocities even with the inclusion of a large loss in the relay time. To this end, we propose a new axon equivalent circuit that incorporates the effect of the dielectric characteristics of the axonal fluid. It was confirmed that a realistically high conduction velocity could be calculated using the proposed circuit and that the dielectric constant calculated using the proposed circuit was in agreement with that of an ionic fluid similar to axonal fluid. Moreover, the contribution of the combination of the axonal morphology and axonal fluid to the conduction velocity was confirmed.

Coordinated peripheral neuronal activities among the different regions of the digestive tract in Aplysia.

Peripheral neuronal somata are scattered throughout the enteric nervous system (ENS) in Aplysia. We found that somata on the outer surface of the digestive tract were more densely distributed on the stomatogastric ring and the posterior gizzard than on other regions. In preparations with or without the central nervous system, two types of synchronous bursting activity were recorded from the nerves of the ENS. Some of the synchronous bursts were recorded from nerves on the crop and stomatogastric ring, whereas others were recorded from nerves on the crop, stomatogastric ring, and gizzard. Experiments using preparations in which the different regions were separated suggested that the former bursts originated in neurons on the crop and the latter originated in neurons on the gizzard. Axonal projections of neurons on the different regions were examined by backfilling and analysis of the direction of impulse conduction. Blocking chemical synapses in separated gizzards depressed EPSP-like potentials and eliminated the bursting activities. When chemical synapses on the crop and stomatogastric ring but not on the gizzard were blocked in a whole digestive tract preparation, bursting activity recorded from nerves on all the regions was decreased, although the frequency of the bursting rhythm did not change. Stimulation of a neuron on the crop elicited bursts in nerves on the gizzard. These results suggest that chemical synaptic connections and a feedback loop along the digestive tract coordinate the synchrony of bursting activity originating in the gizzard.

The main occluding area in normal occlusion and mandibular prognathism.

OBJECTIVE: To clarify whether the concept of main occluding area, where hard food is initially crushed, exists in patients who have a jaw deformity. MATERIALS AND METHODS: Nineteen subjects with normal occlusion, 18 patients with mandibular prognathism, and 11 patients with mandibular prognathism who had undergone orthognathic surgery participated in this study. The main occluding area was identified by clenching Temporary Stopping. The coincidence, location of the main occluding area, and distance from the first molars to main occluding area were examined. RESULTS: High coincidence of the main occluding area was obtained in all groups, signifying that the main occluding area exists even in these patients. Mandibular main occluding area was located on the first molar in all groups. Maxillary main occluding area in subjects with normal occlusion was located on the first molar. However, it was located on the second premolar and first molar in patients with mandibular prognathism, and on the first and second molars in patients with mandibular prognathism who had undergone orthognathic surgery. There was a statistically significant difference in distance from the maxillary first molar to the main occluding area among groups, but there was no difference in the distance from the mandibular first molar among groups. CONCLUSION: The main occluding area is more stable on the mandibular first molar than the maxilla in all groups.

Proton feedback mediates the cascade of color-opponent signals onto H3 horizontal cells in goldfish retina.

It has been postulated that horizontal cells (HCs) send feedback signals onto cones via a proton feedback mechanism, which generates the center-surround receptive field of bipolar cells, and color-opponent signals in many non-mammalian vertebrates. Here we used a strong pH buffer, HEPES, to reduce extracellular proton concentration changes and so determine whether protons mediate color-opponent signals in goldfish H3 (triphasic) HCs. Superfusion with 10mM HEPES-fortified saline elicited depolarization of H3 HCs' dark membrane potential and enhanced hyperpolarizing responses to blue stimuli, but suppressed both depolarization by yellow and orange and hyperpolarization by red stimuli. The response components suppressed by HEPES resembled the inverse of spectral responses of H2 (biphasic) HCs. These results are consistent with the Stell-Lightfoot cascade model, in which the HEPES-suppressed component of H3 HCs was calculated using light responses recorded experimentally in H1 (monophasic) and H2 HCs. Selective suppression of long- or long-+middle-wavelength cone signals by long-wavelength background enhanced the responses to short-wavelength stimuli. These results suggest that HEPES inhibited color opponent signals in H3 HCs, in which the source of opponent-color signals is primarily a feedback from H2 HCs and partly from H1 HCs onto short-wavelength cones, probably mediated by protons.

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 Co(2+) and Ni(2+) 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.

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.