Observational study of effects of pharyngeal stimulation by carbonated solution on repetitive voluntary swallowing in humans.

In this study, we conducted observational study to examine the effects of pharyngeal stimulation by a bolus of carbonated solution on repetitive voluntary swallowing in humans. Twelve healthy participants had a fine silicone tube inserted into their pharyngeal region, through which various solutions were slowly infused (0.2 mL/minute) to stimulate the pharyngeal mucosa without activating mechanoreceptors. The solutions included 0.3M NaCl (NaCl), carbonated 0.3M NaCl (NaCl + CA), 0.3M NaCl with acetic acid, distilled water, and carbonated distilled water. We used NaCl to inhibit water-sensitive neurons in the pharyngeal mucosa and enable the evaluation of the effects of carbonic acid stimulation on swallowing. Participants were instructed to repeat swallows as rapidly as possible during the infusion, and the swallowing interval (SI) was measured via submental surface electromyographic activity. SI was significantly shorter during the infusion of NaCl + CA, distilled water, and carbonated distilled water than during the infusion of NaCl. There was a significant positive correlation between SI with NaCl stimulation and the facilitative effects of the other solutions. Longer SIs with NaCl stimulation indicated potent facilitative effects. Thus, stimulation with NaCl + CA facilitated swallowing by reducing SI. Furthermore, the facilitative effects of SI were more pronounced in participants who had difficulty with repetitive voluntary swallowing. The sensation induced by carbonated solution may enhance the ability for repetitive voluntary swallowing, making it a potentially useful approach for rehabilitating patients with dysphagia.

Facilitation of voluntary swallowing by chemical stimulation of the posterior tongue and pharyngeal region in humans.

In this study, we investigated the functional difference between chemical stimulations of the posterior tongue (PT) and pharyngeal region (PR) for facilitation of voluntary swallowing in humans. The PT or PR stimulation consisted of infusion of water (distilled water), 0.3 M NaCl solution or olive oil (non-chemical stimulant) into the PT or the PR through a fine tube at a very slow infusion rate (0.2 ml/min). Water was used as a stimulant of water receptors. A solution of 0.3 M NaCl was used as an inhibitor of the response of water receptors and as a stimulant of salt taste receptors. Excitation of the mucosal receptors would facilitate voluntary swallowing and diminution of sensory inputs from the oral mucosa would induce difficulty in swallowing. Swallowing intervals (SIs) during voluntary swallowing were measured by submental electromyographic activity. Infusion of water into the PR shortened SI (facilitation of swallowing) and infusion of 0.3 M NaCl or olive oil into the same region prolonged it (difficulty in swallowing). On the other hand, infusion of water into the PT prolonged SI and infusion of 0.3 M NaCl into the same region shortened it. The results suggest that water receptors are localized in the PR and that salt taste receptors are almost absent in the PR and present in the PT. With diminution of sensory inputs from the oral mucosa, central inputs would play a dominant role in initiating swallowing voluntarily, and SI would be prolonged. With weak stimulation (infusion of 0.3 M NaCl into the PR or infusion of water into the PT), SI was prolonged and inter-individual variation in SI was pronounced, suggesting that the ability of the central regulation of swallowing to perform repetitive voluntary swallowing varies among subjects. With stimulation of water receptors or salt taste receptors, SI was shortened and inter-individual variation in SI was moderate, suggesting that sensory inputs are important for performing voluntary swallowing smoothly and that the sensory inputs compensate for the difficulty in performing swallowing caused by the central mechanism.

Mechanism of enhancement of the responses of the frog glossopharyngeal nerve to electrolytes by enhancers.

In frogs, the responses of the glossopharyngeal nerve (GL) to NaCl are enhanced after treatment of the tongue with 8-anilino-1-naphthalene-sulfonic acid (ANS), a hydrophobic probe for biological membranes. The enhancement by ANS treatment has been explained by removal of Ca2+ from the receptor membrane treated with ANS. To explore the mechanism of enhancement by ANS treatment, we recorded neural responses from the frog GL. After ANS treatment, treatment with 10 mM CaCl2 prior to stimulation of NaCl did not affect the enhanced responses to 100 mM NaCl. The response to a relatively high concentration of CaCl2 (50 mM) was enhanced after ANS treatment. It is difficult to interpret these neural events in terms of modulation of the responses by membrane-bound calcium. The presence of NiCl2 in stimulating solution is known as an enhancer. Neural events after ANS treatment were similar to those caused by NiCl2. Our previous studies have demonstrated that enhancement of the responses to electrolytes by NiCl2 is due to modulation of the responses of water fibers in the GL. Water fibers are characterized by sensitivity to water or CaCl2, and they also respond to relatively high concentrations of electrolytes such as NaCl and choline Cl. Using a suction electrode method, we recorded unitary impulses from single water fibers. The ANS treatment led greatly enhanced responses to NaCl or choline Cl in water fibers, suggesting that enhancement by the ANS treatment is due to modulation of the responses of water fibers as well as enhancement by NiCl2. It appears that distinct receptors for each separate cation responsible for the neural responses in water fibers interact with a membrane element that is affected by ANS or Ni2+.

Synaptic responses of neurons controlling the parotid and von Ebner salivary glands in rats to stimulation of the solitary nucleus and tract.

Salivary secretion results from reflex stimulation of autonomic neurons via afferent sensory information relayed to neurons in the rostral nucleus of the solitary tract (rNST), which synapse with autonomic neurons of the salivatory nuclei. We investigated the synaptic properties of the afferent sensory connection to neurons in the inferior salivatory nucleus (ISN) controlling the parotid and von Ebner salivary glands. Mean synaptic latency recorded from parotid gland neurons was significantly shorter than von Ebner gland neurons. Superfusion of GABA and glycine resulted in a concentration-dependent membrane hyperpolarization. Use of glutamate receptor antagonists indicated that both AMPA and N-methyl-D-aspartate (NMDA) receptors are involved in the evoked excitatory postsynaptic potentials (EPSPs). Inhibitory postsynaptic potential (IPSP) amplitude increased with higher intensity ST stimulation. Addition of the glycine antagonist strychnine did not affect the amplitude of the IPSPs significantly. The GABA(A) receptor antagonist, bicuculline (BMI) or mixture of strychnine and BMI abolished the IPSPs in all neurons. IPSP latency was longer than EPSP latency, suggesting that more than one synapse is involved in the inhibitory pathway. Results show that ISN neurons receive both excitatory and inhibitory afferent input mediated by glutamate and GABA respectively. The ISN neuron response to glycine probably derives from descending connections. Difference in the synaptic characteristics of ISN neurons controlling the parotid and von Ebner glands may relate to the different function of these two glands.

Membrane excitability of wing and rod cells in frog taste discs following denervation.

The frog tongue has a disc-shaped taste organ (taste disc) on the top of fungiform papillae. The taste disc contains two types of cells, wing cells with a sheet-like apical process and rod cells with a rod-like apical process. Both wing and rod cells can produce action potentials. Unlike the taste buds of mammals, frog taste discs do not degenerate over a long period after denervation. Here we report that the shapes of wing and rod cells isolated from taste discs in the bullfrog (Rana catesbeiana) remained unchanged 1 month after cutting bilateral glossopharyngeal nerves. By applying the whole cell patch-clamp technique to isolated wing and rod cells, we found voltage-dependent inward currents and outward currents and action potentials in denervated frogs as seen in normal frogs. These results suggest that the maintenance of morphological integrity and electrical excitability of taste cells does not require a nerve supply in frogs.

Bilateral projection of functionally characterized trigeminal oralis neurons to trigeminal motoneurons in cats.

Intracellular Neurobiotin-injections were used to label functionally identified neurons in the rostro-dorsomedial part of the trigeminal oral nucleus (Vo.r) in the cat. The labeled Vo.r neurons with the mechanoreceptive field in oral tissues innervated bilaterally either jaw-opening motoneurons or jaw-closing motoneurons. This result suggests that Vo.r neurons play an important role in sensory-motor reflexes responsible for coordination of bilaterally symmetrical jaw movements.

Biophysical and morphological properties of parasympathetic neurons controlling the parotid and von Ebner salivary glands in rats.

The inferior salivatory nucleus (ISN) contains parasympathetic neurons controlling the parotid and von Ebner salivary glands. To characterize the neurophysiological and morphological properties of these neurons, intracellular recordings were made from anatomically identified ISN neurons in rat brain slices. Neurons were also filled with Lucifer yellow and morphometrically analyzed. Based on responses to membrane hyperpolarization followed by depolarization, three types of repetitive discharge patterns were defined for neurons innervating the parotid gland. The regular, repetitive discharge response to membrane depolarization was changed by hyperpolarization resulting either in a delay in the occurrence of the first spike or to an increase in the length of the first interspike interval in the action potential train. Membrane hyperpolarization had little effect on the discharge pattern of some neurons. Similar response discharge patterns were found for neurons innervating the von Ebner salivary gland, which also included a further group of neurons that responded with a short burst of action potentials. Neurons innervating the parotid salivary glands differed morphologically from the von Ebner salivary glands having significantly larger soma and more and longer dendrites than von Ebner gland neurons. In addition, the mean membrane input resistance, time constant, and spike half-width of parotid gland neurons was significantly lower than in von Ebner gland neurons. These differences in intrinsic membrane properties and morphology may relate to the functions of the von Ebner and parotid glands. von Ebner glands are involved in taste stimulus delivery and removal from posterior tongue papillae while the parotid glands contribute saliva to the entire mouth.

Quantitative analysis of the dendritic architectures of single jaw-closing and jaw-opening motoneurons in cats.

Little is known about the dendritic architectures of trigeminal motoneurons innervating antagonistic muscles. Thus, the aim of the present study was to provide a quantitative description of jaw-closing (JC) and jaw-opening (JO) alpha motoneurons and to determine geometrical similarities and differences of the dendritic tree between the two. Seven JC alpha motoneurons and four JO alpha motoneurons were intracellularly labeled with horseradish peroxidase (HRP) in the cat and quantitatively analyzed with a computer-assisted three-dimensional system. The dendritic tree of JC alpha motoneurons was confined within the JC motor nucleus, despite locations of the cell body. In contrast, JO alpha motoneurons generated extensive extranuclear dendrites in the reticular formation. The branching pattern of proximal dendritic segments was simpler in the JC than in the JO alpha motoneurons. Despite these differences, the mean values of dendritic parameters examined per neuron were not different between the two kinds of alpha motoneurons, and the stem dendrite diameter was positively correlated with several dendritic parameters in a linear manner. The present study provides new evidence that underlying design principles of the geometry of the dendritic tree are not concerned with the differences in configuration and branching pattern of the dendritic tree of trigeminal alpha motoneurons innervating antagonistic muscles. In addition, we estimated the number of excitatory and inhibitory synapses covering dendrites of single JC alpha motoneurons.

NADPH-diaphorase and calcium binding proteins in the trigeminal nucleus oralis of rats.

We have examined the distribution of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and the calcium binding proteins (CBPs), calbindin D-28k (CB), calretinin (CR) and parvalbumin (PV), in the trigeminal nucleus oralis (Sp5O). NADPH-d was detected by histochemistry while CBP was detected by immunohistochemistry. NADPH-d-positive neurons were distributed in the medial rostro-dorsomedial part (RDMsp5O) and dorsomedial part (DMsp5O) of Sp5O, and the rostrolateral part of the nucleus of the solitary tract (NTS). CB- and CR-positive neurons were mainly distributed in the dorsal part of Sp5O. In contrast, PV-positive neurons were mainly distributed in the ventral part of Sp5O. NADPH-d colocalized with CB (40%) and CR (20%) but not with PV in neurons of DMsp5O/ NTS. The mean cell sizes of neurons in RDMsp5O were larger than those in DMsp5O/NTS. PV-positive neurons were larger than NADPH-d-positive neurons. NADPH-d-, CB- and CR-positive neurons were generally small in RDMsp5O and DMsp5O/NTS. Few neurons were retrogradely labeled in RDMsp5O and DMsp5O from the thalamus, when numerous labeled neurons were in the principal and interpolar nuclei. These data indicate that NADPH-d histochemistry and CB, CR and PV immunohistochemistry identify a discrete cell population in Sp5O. Those labeled neurons in RDMsp5O and DMsp5O/NTS were considered to be involved in sensorimotor reflexive function of the intra-oral structures.

Quantitative ultrastructural analysis of glycine- and gamma-aminobutyric acid-immunoreactive terminals on trigeminal alpha- and gamma-motoneuron somata in the rat.

Detailed knowledge of the inhibitory input to trigeminal motoneurons is needed to understand better the central mechanisms of jaw movements. Here a quantitative analysis of terminals contacting somata of jaw-closing (JC) and jaw-opening (JO) alpha-motoneurons, and of JC gamma-motoneurons, was performed by use of serial sectioning and postembedding immunogold cytochemistry. For each type of motoneuron, the synaptic boutons were classified into four groups, i.e., immunonegative boutons or boutons immunoreactive to glycine only, to gamma-aminobutyric acid (GABA) only, or to both glycine and GABA. The density of immunolabeled boutons was much higher for the alpha- than for the gamma-motoneurons. In the alpha-motoneuron populations, the immunolabeled boutons were subdivided into one large group of boutons containing glycine-like immunoreactivity only, one group of intermediate size harboring both glycine- and GABA-like immunoreactivity, and a small group of boutons containing GABA-like immunoreactivity only. The percentage of immunolabeled boutons was higher for JC than JO alpha-motoneurons, the most pronounced difference being observed for glycine-like immunoreactivity. In contrast, on the somatic membrane of gamma-motoneurons, the three types of immunoreactive bouton occurred at similar frequencies. These results indicate that trigeminal motoneurons are strongly and differentially controlled by premotoneurons containing glycine and/or GABA and suggest that these neurons play an important role for the generation of masticatory patterns.