Fusako Kagitani

Effect of stimulation of nicotinic cholinergic receptors on cortical cerebral blood flow and changes in the effect during aging in anesthetized rats

The effect of intravenous injection of nicotine on cortical cerebral blood flow (CBF) was examined in urethane anesthetized rats. Nicotine (3-30 microg/kg) increased cortical CBF, independent of mean arterial pressure. This response was attenuated to about a half of the control one after lesioning the nucleus basalis of Meynert (NBM) bilaterally. The response was not significantly influenced after blocking the muscarinic receptors, but was abolished after blocking the nicotinic receptors in the parenchyma of the brain. It is concluded that the nicotine-induced cortical vasodilation was mediated by activation of the nicotinic receptors in the NBM and also in the cortex of the brain. The threshold dose of nicotine for increasing cortical CBF was shifted in aged rats of 23-26 months, and the nicotine-induced increase in cortical CBF was much reduced in aged rats of 32-36 months. Activation of nicotinic receptors in the brain may be of therapeutic value in aged subjects in facilitating the cholinergic neural vasodilative system.

Afferent nerve fibers and acupuncture

Acupuncture has been used for analgesia, for treating visceral function disorders and for improving motor functions. It is well established that stimulation of the skin and muscles, either electrically or with noxious or non-noxious stimuli, induces a variety of somato-motor and autonomic responses. This strongly suggests that acupuncture acts by exciting cutaneous and/or muscular afferent nerve fibers. A question of considerable scientific and practical interest is what kinds of somatic afferent fibers are stimulated by acupuncture and are involved in its effects. There are several types of afferent fiber: thick myelinated Aα and Aβ (group I and II), thin myelinated Aδ (group III) and thinner unmyelinated C (group IV) fibers. In recent studies we have tried to establish which ones of these types of somatic afferent fiber are stimulated by acupuncture. In this article we first review the experimental evidence showing that the effects of acupuncture are mediated by the activation of afferent nerve fibers innervating the skin and muscles. Secondly, we discuss what types of afferent nerve fiber are activated by electrical acupuncture, and what types are involved in its effects on somato-motor functions and on visceral functions. Finally, we present some new findings based on recordings from single afferent nerve fibers excited by manual acupuncture.

Effects of age on cholinergic vasodilation of cortical cerebral blood vessels in rats

The present study examined the age-related changes in the cholinergic vasodilative system originating in the nucleus basalis of Meynert (NBM) and projecting to the cerebral cortex using Wistar rats of three different ages; young adult (4-7 months), old (24-25 months), and very old (32-42 months) rats. The vasodilative responses in frontal and parietal cortices, measured by laser Doppler flowmetry, induced by electrical stimulation of NBM without blood pressure response were well maintained in old rats, but declined significantly in very old rats. Extracellular acethylcholine (ACh) release in both cortices collected by a microdialysis technique showed both basal levels and response to NBM stimulation to be well maintained in both old and very old rats. The vasodilative cerebral blood flow response elicited by stimulation of the muscarinic ACh receptors, using their agonist, arecoline, was also well maintained in old and very old rats. Considering the present data and our previous finding that the cerebral cortical vasodilative response to activation of the nicotinic ACh receptors using their agonist, nicotine, was markedly reduced in very old rats (Neurosci. Lett., 228 (1997) 203), it was concluded that the age-related decline of nicotinic ACh receptor activity was a cause of the decline of the vasodilative responses elicited by NBM stimulation in very old rats. This result suggests that a reduction of the cholinergic vasodilative system in very old rats due to decreased activity of the nicotinic ACh receptor may cause insufficient blood flow in the cortex when the cortical neurons require.

Basal forebrain stimulation induces NGF secretion in ipsilateral parietal cortex via nicotinic receptor activation in adult, but not aged rats

The role of cholinergic basal forebrain inputs to cerebral cortex in regulating regional nerve growth factor (NGF) secretion was examined in adult (4-6 months) and aged (29-31 months) rats. Halothane-anesthetized rats received unilateral electrical stimulation of the nucleus basalis of Meynert (NBM). NGF levels were measured by ELISA in samples from a microdialysis probe in the parietal cortex, while measuring blood flow. In adult and aged rats, NBM stimulation (for 100 min) increased blood flow ipsilaterally during stimulation by 55% and 25%, respectively. In adult, but not aged rats, NGF levels were significantly increased ipsilaterally (up to 68%) over prestimulus levels at 200-500 min after stimulation ended. The cellular localization of NGF-like immunoreactivity showed no differences between the cortices with and without NBM stimulation. The NGF response was abolished by the nicotinic blocker, mecamylamine (20 mg/kg iv), but unaffected by the muscarinic blocker, atropine (5 or 25 mg/kg iv). Both drugs reduced the blood flow responses. We conclude that cholinergic inputs to neocortex mediate NGF secretion by cortical neurons via nicotinic receptors. Further, the absence of this response in aged rats suggests a decline in the number or activity of cortical nicotinic receptors.

Mechanism of the reflex inhibition of heart rate elicited by acupuncture-like stimulation in anesthetized rats

Acupuncture or acupuncture-like stimulation applied to different body areas can modify autonomic nerve activity to various organs, including gut, bladder, adrenal medulla, and the heart. We studied the reflex bradycardia in response to insertion into the skin and underlying muscles and twisting of an acupuncture needle in pentobarbital-anesthetized rats. We found that acupuncture-like stimulation of forelimb, hindlimb, chest, and abdomen all produced significant heart rate decreases. Rate minima were reached at the end of the 60-second stimulation episode and significant bradycardia persisted for about 40 s after stimulation ended. Heart rate decreases were paralleled by decreases in cardiac sympathetic nerve activity, and could be produced by electrical stimulation of group IV muscle afferent fibers (tibial nerve). Electrical stimulation of the tibial nerve at rates as low as 0.1-2 Hz was effective for eliciting heart rate decreases. Nerve fiber groups were defined by stimulation of and recording from tibial nerve. Activation of groups I, II, or III fiber was ineffective for eliciting the reflex bradycardia. Sympathectomy, high spinal transection, or infusion of the GABA(A) receptor antagonist, bicuculline, into the cisterna magna were all effective for disrupting the reflex bradycardia. Vagotomy and opioid receptor blockade were ineffective for disrupting the reflex pathway. We conclude that the reflex pathway to decrease heart rate by acupuncture-like stimulation consists of mainly group IV muscle afferent fibers whose activity (even very low rate of activity) leads to the activation of GABA-ergic neurons in the brainstem and an inhibition of sympathetic outflow to the heart.

Cilostazol, a selective cAMP phosphodiesterase inhibitor, dilates retinal arterioles and increases retinal and choroidal blood flow in rats

The effects of cilostazol, a selective cyclic AMP (cAMP) phosphodiesterase inhibitor, on retinal and choroidal blood flow and retinal arteriole diameter were examined in anesthetized rats. The retinal and choroidal blood flow was measured using laser Doppler flowmetry, and the diameter of the retinal arterioles was measured using digital video microscopy. Cilostazol was administered by two routes; systemically via the intravenous route, and directly into the retinal vessels via the intra-arterial route. When administered intravenously, 1 mg/kg of cilostazol produced a biphasic blood flow response, composed of an initial decrease which was dependent on a depressor response of mean arterial pressure, and a subsequent slight but significant increase which was independent of changes in mean arterial pressure. When administered intra-arterially over a 2-min period, 40-55 and 400-440 microg of cilostazol both produced an increase in the blood flow in a dose-dependent manner, while a depressor effect was observed only at the dose of 400-440 microg. The diameter of the retinal arterioles was increased after the intra-arterial injection of cilostazol (400 microg). It is concluded that intra-arterially administered cilostazol induces vasodilation of the retinal arterioles of rats, which results in an increase in blood supply to the retina, independent of changes in mean arterial pressure.

Effect of Moxibustion Stimulation of Various Skin Areas on Cortical Cerebral Blood Flow in Anesthetized Rats

The effect of moxibustion stimulation of various skin areas (cheek, forepaw, upper arm, chest, back, lower leg, hindpaw and perineum) on cerebral blood flow (CBF) of the parietal cortex was examined in anesthetized rats after eliminating emotional influences. Moxibustion stimulation was performed by burning a moxa cone of about 4 mg weight placed on the shaved skin. CBF of the parietal cortex was measured using a laser Doppler flowmeter. Stimulation of the cheek, forepaw, upper arm and hindpaw produced significant increases in CBF, but stimulation of the other areas did not produce significant responses. Moxibustion stimulation of the forepaw and hindpaw produced an increase in the mean arterial pressure (MAP), while stimulation of the other areas did not. After spinal transection at the 2nd thoracic level, the MAP response to stimulation of the forepaw was abolished, whereas the CBF response to stimulation of the forepaw remained. The CBF response in spinalized rats was not affected by cutting cervical sympathetic and facial parasympathetic nerves, while it was almost abolished by intravenous administration of muscarinic and nicotinic cholinergic blocking agents. The CBF response was abolished by crushing the brachial plexus ipsilateral to the stimulated side. It is suggested that the increase in CBF, independent of MAP and emotional responses, elicited by moxibustion stimulation is a reflex response whose afferent pathway is composed of somatic afferent nerves, and whose efferent pathway involves intracerebral cholinergic nerves. A contribution of endogenous opioids in the present CBF responses was neglected, because naloxone did not influence the CBF responses.

Effects of Electrical Stimulation of the Superior Ovarian Nerve and the Ovarian Plexus Nerve on the Ovarian Estradiol Secretion Rate in Rats

The present experiments examined the effects of electrical stimulation of the superior ovarian nerve (SON) and the ovarian plexus nerve (OPN) on the ovarian estradiol secretion in rats. The rats were anesthetized on the day of estrus, and the ovarian venous blood was collected intermittently. The secretion rate of estradiol from the ovary was calculated from differences in the estradiol concentration between ovarian venous plasma and systemic arterial blood plasma, and from the flow rate of ovarian venous plasma. Either an SON or OPN, ipsilateral to the ovary from which ovarian venous blood was collected, was electrically stimulated at a supramaximal intensity for C-fibers. The secretion rate of estradiol was significantly decreased by 47 +/- 6% during SON stimulation, but it was not significantly changed during OPN stimulation. These results suggest that autonomic nerves, which reach the ovary via the SON, have an inhibitory role in ovarian estradiol secretion.

Neural mechanisms of reflex inhibition of heart rate elicited by acupuncture-like stimulation in anesthetized rats

We briefly review our recent studies on the neural mechanisms of the reflex effects of acupuncture-like stimulation on heart rate in rats. In pentobarbital anesthetized rats, acupuncture-like stimulation of one of various segmental areas of the body (forelimb, chest, abdomen, hindlimb) invariably induces a decrease in heart rate. In the case of the hindlimb, the effect can be produced by stimulation of the muscles alone but not of skin alone, and is abolished by severance of the hindlimb somatic nerves. Electrical stimulation of groups III and IV nerve fibers (in the tibial nerve) decreases heart rate. Decrease in heart rate by acupuncture-like stimulation of a hindlimb is accompanied by a decrease in cardiac sympathetic nerve activity, and is abolished by cardiac sympathectomy but not by vagotomy. High spinal cord transection or infusion of the GABA(A) receptors antagonist, bicuculline, into the cisterna magna is effective in disrupting the reflex bradycardia. Opioid receptor blockade does not disrupt the reflex arc. We conclude that the reflex pathway involved in the decrease of heart rate by acupuncture-like stimulation comprises groups III and IV muscle afferent nerves whose activation stimulates GABAergic neurons in the brainstem and inhibits sympathetic outflow to the heart. When the sympathetic tone is high due to hypercapnia, the induced reduction in both cardiac sympathetic nerve activity and heart rate is not augmented, suggesting that the magnitude of sympatho-inhibitory response to acupuncture-like stimulation does not depend on pre-existing sympathetic tone.

Control of cerebral cortical blood flow by stimulation of basal forebrain cholinergic areas in mice

We examined whether activity of the nucleus basalis of Meynert (NBM) regulates regional cerebral cortical blood flow (rCBF) in mice, using laser speckle and laser Doppler flowmetry. In anesthetized mice, unilateral focal stimulation, either electrical or chemical, of the NBM increased rCBF of the ipsilateral cerebral cortex in the frontal, parietal and occipital lobes, independent of changes in systemic blood pressure. Most of vasodilative responses to low intensity stimuli (2 times threshold intensity: 2T) were abolished by atropine (a muscarinic cholinergic blocker), whereas responses to higher intensity stimuli (3T) were abolished by atropine and mecamylamine (a nicotinic cholinergic blocker). Blood flow changes were largest when the tip of the electrode was located within the area containing cholinergic neurons shown by choline acetyltransferase-immunocytochemistry. These results suggest that cholinergic projections from basal forebrain neurons in mice cause vasodilation in the ipsilateral cerebral cortex by a combination of muscarinic and nicotinic mechanisms, as previously found in rats and cats.