Hisayoshi Ishii

Evidence for parasympathetic vasodilator fibres in the rat masseter muscle

The present study was designed to examine (1) whether there are vasodilator fibres in the masseter muscle, and (2) if there are, to establish the neural pathways mediating these responses in urethane‐anaesthetized rats. Electrical stimulation of the central cut end of the lingual nerve (LN) elicited intensity‐ and frequency‐dependent increases of the blood flow in the masseter muscle (MBF) and lower lip (LBF). Increases in both the MBF and LBF evoked by the LN stimulation were reduced by hexamethonium in a dose‐dependent manner (1–10 mg kg−1). Pretreatment with phentolamine or propranolol at a dose of 100 μg kg−1 had no effect on the increases in either MBF or LBF evoked by LN stimulation. Pretreatment with atropine (100 μg kg−1) significantly reduced the MBF increase induced by LN stimulation, but not that in the LBF. The sectioning of the superior cervical sympathetic trunk did not affect the responses. MBF increases occurred with electrical stimulation of the trigeminal ganglion, and these increases were significantly reduced by the administration of hexamethonium and atropine. Lidocaine microinjection into the trigeminal spinal nucleus or salivatory nuclei caused a significant attenuation of the LN‐induced MBF increases. When wheat germ agglutinin–horseradish peroxidase (WGA–HRP) was injected into the masseter muscle, labelled neurones were abundantly observed in the otic ganglion. The present study indicates that there are parasympathetic cholinergic and noncholinergic vasodilator fibres originating from cell bodies in the otic ganglion in the rat masseter muscle. The MBF increase evoked by activation of the parasympathetic fibres occurred via the trigeminal mediated reflex, suggesting that the novel parasympathetic vasodilator response may play an important role in the regulation of the haemodynamics of jaw muscles.

Vagal visceral inputs to the nucleus of the solitary tract: Involvement in a parasympathetic reflex vasodilator pathway in the rat masseter muscle

The present study examined whether vagal visceral inputs are involved in parasympathetic reflex vasodilatation in the masseter muscle in urethane-anesthetized and cervically sympathectomized rats. Electrical stimulation of the central cut end of the cervical vagus nerve (VN) including visceral afferent fibers, which consisted of cervical/thoracic branches (heart and lungs) and abdominal branches (entire gastrointestinal tract), elicited intensity- and frequency-dependent increases of blood flow in the masseter muscle (MBF). Activation of the abdominal VN inferior to the diaphragm failed to affect the MBF. MBF increases evoked by cervical VN stimulation were reduced significantly by hexamethonium. Pretreatment with atropine reduced the MBF increase evoked by VN stimulation significantly, whereas pretreatment with either propranolol or phentolamine had no effect on the response. MBF increases occurred with electrical stimulation of nucleus of the solitary tract (NTS), and these increases were significantly reduced by the administration of hexamethonium and atropine. MBF increases also occurred after microinjection of glutamate into the NTS in a dose-dependent manner. Microinjection of muscimol into the NTS caused a significant attenuation of the VN stimulation-induced MBF increases. Our results suggest that vagal visceral inputs passing to the NTS are involved in the parasympathetic reflex vasodilatation in the rat masseter muscle. The MBF increase evoked by the vagal-parasympathetic reflex mechanism occurred via visceral afferents running in the cervical VN, but not in the abdominal VN, suggesting that the vagal visceral afferents derived from cardiovascular and/or respiratory systems may play an important role in the regulation of the MBF.

Differential gene expression of vascular endothelial growth factor isoforms and their receptors in the development of the rat masseter muscle.

The capillary network in the masseter muscle develops dramatically with the differentiation of muscle fibres after birth, especially around weaning. Here, developmental changes in mRNA expression for four splicing variants of vascular endothelial growth factor (VEGF) and for two distinct VEGF receptors (Fms-like tyrosine kinase (Flt-1) and kinase insert domain-containing receptor/fetal liver kinase-1 (KDR/Flk-1)) were studied in rat masseter. The relative abundance of VEGF (120) mRNA was the highest, representing 35% of total VEGF mRNA on day 7 after birth and gradually decreased with age to become approximately 5% on day 37. In contrast, VEGF (188) mRNA was very low in the newborn rat, but increased sharply before weaning and reached 40-50% of the total on day 50. Neither VEGF (144) nor VEGF(164) mRNA showed any significant change in abundance after birth. The expression of KDR/Flk-1 mRNA was transiently high in the early postnatal stage and gradually decreased with age, Flt-1 mRNA was stably expressed at a constant level after birth. These findings suggest that different combinations of VEGF isoforms and their receptors regulate angiogenesis in the development of the masseter muscle.

Difference between male and female rats in cholinergic activity of parasympathetic vasodilatation in the masseter muscle

We compared the changes in blood flow of the masseter muscle (MBF), lower lip (LBF) and common carotid artery (CCABF) evoked by electrical stimulation of the lingual nerve (LN) in order to examine whether high cholinergic activity of parasympathetic vasodilatation in females is specific for the masseter muscle, and whether sex-associated differences in cholinergic parasympathetic vasodilatation affect the regulation of blood flow to the orofacial area from the CCABF in urethane-anaesthetized, vago-sympathectomized male and female rats. Increases in the MBF, LBF and CCABF evoked by LN stimulation appear to be mediated via an activation of parasympathetic reflex vasodilatation since these increases were profoundly reduced by pretreatment with the autonomic cholinergic ganglion blocker hexamethonium (10 mg/kg). Although alpha- and beta-adrenoceptor antagonists (phentolamine and propranolol, 100 microg/kg) had no effect on the LN stimulation-induced blood flow increases in either sex, a marked difference was found between males and females in the effects of the antimuscarinic agent atropine (1-100 microg/kg) on these blood flow increases. Pretreatment with atropine slightly attenuated the increase in the MBF in males, but in females it markedly reduced the increases in all three sites measured, especially in the MBF. Our results suggest that (1) cholinergic activity of the parasympathetic vasodilatation in females is higher than that in males in most orofacial tissues, but particularly in the masseter muscle and (2) cholinergic parasympathetic vasodilatations are more involved in the regulation of blood flow to the orofacial area from the CCABF in females than in males.

Circulating adrenaline released by sympathoadrenal activation elicits acute vasodilatation in the rat masseter muscle

The present study was designed to examine the effects of circulating catecholamines released by sympathoadrenal system on the haemodynamics of the masseter muscle in deeply urethane-anaesthetized, artificially ventilated, cervically vagotomized and sympathectomized rats. Intravenous administration of adrenaline induced a biphasic change of blood flow in the masseter muscle (MBF). The change of blood flow showed an initial marked increase and successive slight decrease in a dose-dependent manner (0.01-1 microg/kg). The administration of noradrenaline had no significant effect on the MBF. The increase in the MBF evoked by exogenously applied adrenaline was markedly reduced by the intravenous administration of propranolol (100 microg/kg), whereas pretreatment with either hexamethonium (10 mg/kg), atropine (100 microg/kg), or phentolamine (1 mg/kg) failed to affect the MBF increase. Electrical stimulation of splanchnic nerve (SPLN) preganglionic neurones projecting to the adrenal medulla elicited frequency-dependent (1-20 Hz) increases in the MBF. The intravenous administration of the beta(2)-adrenergic receptor selective antagonist, ICI 118551 (0.5 mg/kg), almost abolished the MBF increase induced by SPLN stimulation, but pretreatment with the beta(1)-adrenergic receptor selective antagonist, atenolol (1 mg/kg), had no effect on this response. The results of the present study indicate that circulating adrenaline elicits acute vasodilatation through a beta-adrenergic mechanism in the rat masseter muscle. Vascular beta(2)-adrenergic receptors in the masseter muscle may be activated preferentially by adrenaline released from the adrenal medulla, suggesting that the sympathoadrenal system is involved in the marked MBF increase during sympathoexcitation.

Regional differences in blood flow variation in rat masseter muscle.

Regions of a belly in a masseter muscle have been suggested to be activated independently in order to enable complex jaw-movements. However a regional difference of the masseter blood flow (MBF) is still unclear although the blood flow is one of the most important factors during activation of the muscle. The present study examined regional differences in blood flow in rat masseter muscle by comparing blood flow values at the inferior, centre, superior, anterior, and posterior regions of the muscle belly using a laser speckle imaging flowmeter with or without sympathetic and parasympathetic stimulation. Regional differences in blood flow levels were observed in each region of the masseter muscle belly during rest. Additionally, amplitudes of blood flow changes evoked by electrical stimulation of parasympathetic and sympathetic nerves differed among regions. These results demonstrate the regional differences in hemodynamics during rest, sympathetic vasoconstriction (including the recovery phase), and parasympathetic vasodilatation in rat masseter muscle.

Developmental expression of vascular endothelial growth factor in the masseter muscle of rats

Developmental changes in vascular endothelial growth factor (VEGF) in rat masseter after birth were investigated. VEGF was extracted efficiently and reproducibly from muscle homogenate with low concentrations of non-ionic detergents, such as Triton X-100, Nonidet P-40, and Tween 20. The amount of VEGF measured by enzyme-linked immunosorbent assay (ELISA) increased markedly by approximately 9-fold, from day 8 to 35 after birth. The increase in VEGF was closely correlated with the development of the capillary network, as shown by the capillary to muscle fibre ratio (C/F ratio). Immunoblotting revealed that the predominant molecular species of VEGF concentrated with heparin-sepharose beads was VEGF(188). These results suggest that VEGF plays an important part in the development and maintenance of the capillary network in the rat masseter.

GABAB receptors in the NTS mediate the inhibitory effect of trigeminal nociceptive inputs on parasympathetic reflex vasodilation in the rat masseter muscle

The present study was designed to examine whether trigeminal nociceptive inputs are involved in the modulation of parasympathetic reflex vasodilation in the jaw muscles. This was accomplished by investigating the effects of noxious stimulation to the orofacial area with capsaicin, and by microinjecting GABA(A) and GABA(B) receptor agonists or antagonists into the nucleus of the solitary tract (NTS), on masseter hemodynamics in urethane-anesthetized rats. Electrical stimulation of the central cut end of the cervical vagus nerve (cVN) in sympathectomized animals bilaterally increased blood flow in the masseter muscle (MBF). Increases in MBF evoked by cVN stimulation were markedly reduced following injection of capsaicin into the anterior tongue in the distribution of the lingual nerve or lower lip, but not when injected into the skin of the dorsum of the foot. Intravenous administration of either phentolamine or propranolol had no effect on the inhibitory effects of capsaicin injection on the increases of MBF evoked by cVN stimulation, which were largely abolished by microinjecting the GABA(B) receptor agonist baclofen into the NTS. Microinjection of the GABA(B) receptor antagonist CGP-35348 into the NTS markedly attenuated the capsaicin-induced inhibition of MBF increase evoked by cVN stimulation, while microinjection of the GABA(A) receptor antagonist bicuculline did not. Our results indicate that trigeminal nociceptive inputs inhibit vagal-parasympathetic reflex vasodilation in the masseter muscle and suggest that the activation of GABA(B) rather than GABA(A) receptors underlies the observed inhibition in the NTS.

Parasympathetic reflex vasodilatation in the masseter muscle compensates for carotid hypoperfusion during the vagus-mediated depressor response

Parasympathetic vasodilatation in the orofacial area is thought to be an important factor in the regulation of blood flow in the common carotid artery (CABF), and disturbances in parasympathetic vasodilatations may be related to impairment of the CABF inducing craniofacial ischemia. We hypothesized that the parasympathetic vasodilatation in the masseter muscle evoked by a vagus-mediated reflex is involved in the maintenance of the CABF during the vagus-mediated depressor response. In the present study, we compared changes in blood flow in the masseter muscle (MBF) and CABF, and systemic arterial blood pressure (SABP) evoked by electrical stimulation of the central cut end of the cervical vagus nerve (cVN) in anesthetized and sympathectomized rats. Electrical stimulation of the cVN in the sympathectomized animals caused an increase in MBF followed by a CABF increase, although it simultaneously induced a decrease in SABP. These increases in blood flow changed to decreases after intravenous administration of atropine (100 μg/kg), while pretreatment with atropine had no effect on the changes in SABP. Microinjection (50 nl/site) of the muscimol (1mM), into the nucleus of the solitary tract, which is involved in reflex cardiovascular regulation, markedly inhibited the cVN stimulation-induced MBF increase. Our results indicate that vagal-parasympathetic vasodilatation in the masseter muscle compensates for carotid hypoperfusion during the vagus-mediated depressor response, and that GABAergic neurons may be involved in the inhibition of this response. This inhibition may result in the impairment of CABF, suggesting an important role in the etiology of neurally mediated syncope.

Parasympathetic vasodilator fibers in rat digastric muscle

The present study examined whether parasympathetic vasodilator fibers exist in rat jaw-opening muscles such as the digastric muscle. The mental nerve was stimulated to activate the parasympathetic vasodilator nerve in the digastric muscle. Electrical stimulation of the mental nerve elicited intensity- and frequency-dependent increases of blood flow in this muscle. These increases were markedly reduced by hexamethonium in dose- and time-dependent manners, but pretreatment with phentolamine or propranolol had no effect. Pretreatment with atropine also attenuated the increase in blood flow in digastric muscle. When retrograde fluorogold was injected into the digastric muscle, labeled neurons were observed in the otic ganglion only on the ipsilateral side, but not in the pterygopalatine ganglion of either side. These results indicate that parasympathetic vasodilator fibers originate from cell bodies in the otic ganglion in rat digastric muscle.