OBJECTIVETo investigate the changes of cholinergic nerves in doxorubicin (DOX)-induced rat failing heart and tumor necrosis factor-α (TNF-α) in the heart tissue and serum.

METHODSAdult Sprague-Dawley rats were randomized into control (n=10) and DOX-induced chronic heart failure (CHF) groups (n=15), and in the latter group, the rats were given intraperitoneal injections of 2.5 mg/kg DOX once a week for 6 weeks, with a total cumulative dose of 15 mg/kg. The control rats were injected with normal saline (1 ml/week). Karnovsky-Roots histochemical staining combined with point counting was used to demonstrate the distribution of cholinergic nerves in the heart. The expression levels of TNF-α in the heart tissue and serum were determined with ELISA.

RESULTSPositively stained cholinergic nerves were found in all the rat hearts in the two groups, but in CHF group, the point counts of cholinergic nerves were significantly lower than that of the control group (P<0.01). Compared with the control rats, those with DOX-induced CHF showed elevated levels of TNF-α both in the heart tissue and in the serum (P<0.01).

CONCLUSIONIn rats with DOX-induced CHF, the parasympathetic nervous system is down-regulated in the failing heart, and the diminished cholinergic anti-inflammatory pathway may play an important role in the progression of CHF.

Animals ; Cholinergic Agents ; pharmacology ; Cholinergic Fibers ; drug effects ; Doxorubicin ; pharmacology ; Heart ; drug effects ; innervation ; Heart Failure ; chemically induced ; metabolism ; Male ; Myocardium ; metabolism ; Rats ; Rats, Sprague-Dawley ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo investigate the effect of total fiavonoids from Chrysanthemun morifolium (TFCM) on learning and memory, and cholinergic system function in aging mice.

METHODSThe aging mice model was established by subcutaneous injection of D-galactose. ICR mice were divided into five groups (n=10): contrA group, model group, and TFCM groups. Mice in TFCM groups were given TFCM (50,100 or 150 mg/kg) by gastric irrigation once a day. Learning and memory ability were evaluated by Morris water maze test. The MDA content, SOD and Ach E activity were also measured.

RESULTSCompared with control group, learning and memory ability declined in the D-galactose-induced aging mice; meanwhile MDA content and AchE activity increased, SOD activity decreased. Treatment with TFCM (100, 150 mg/kg) ameliorated the decrease in learning and memory ability of aging mice. Compared with model group, TFCM (100, 150 mg/kg) could also decrease MDA content and Ach E activity, and increase SOD activity in aging mice.

CONCLUSIONTFCM may improve the learning and memory ability of aging mice. The mechanism is involved in its antioxidative characteristic and improvement of central cholinergic system function.

Aging ; drug effects ; physiology ; Animals ; Antioxidants ; pharmacology ; Cholinergic Fibers ; physiology ; Cholinergic Neurons ; physiology ; Chrysanthemum ; chemistry ; Female ; Flavonoids ; isolation & purification ; pharmacology ; Learning ; drug effects ; Male ; Memory ; drug effects ; Mice ; Mice, Inbred ICR

OBJECTIVETo investigate the relation between the progressive effects of chronic intermittent hypoxia (CIH) on cognitive function and the change of cholinergic neuron.

METHODSForty adult male Sprague-Dawley rats were randomly averagely divided into four groups: control group, CIH 1 week group, CIH 3 week group and CIH 5 week group. The cognitive function was assessed by the Morris Water Maze. The necrosis neurons in prefrontal cortex and hippocampus were observed and counted. The cholin acetyltransferase (ChAT) immunostained cells in prefrontal cortex and hippocampus were identified and quantitated.

RESULTSThe spatial learning and memory impairments progressed from 1 to 5 5 weeks in rats. Compared with the control group, the cognitive impairments in CIH5w group were significant (P < 0.05). The degeneration or necrosis neurons in prefrontal cortex and hippocampus were significantly increased in CIH rats, and worsen gradually along with the hypoxia. The ChAT immunostained cells in prefrontal cortex and hippocampus were gradually reduced. The ChAT immunostained cells of prefrontal cortex and hippocampus in CIH3w group and CIH5w group were less than that in control group (P < 0.05).

CONCLUSIONChronic intermittent hypoxia induced slowly progressive spatial learning and memory impairments in rats, which maybe associated with the damage of neurons and the reduction of ChAT in prefrontal cortex and hippocampus.

Animals ; Cholinergic Fibers ; pathology ; physiology ; Cholinergic Neurons ; pathology ; physiology ; Cognition ; physiology ; Hippocampus ; cytology ; physiopathology ; Hypoxia ; physiopathology ; Male ; Maze Learning ; physiology ; Memory Disorders ; etiology ; physiopathology ; Prefrontal Cortex ; cytology ; physiopathology ; Rats ; Rats, Sprague-Dawley

Parkinson's disease is a progressive neurodegenerative disorder characterized clinically by rigidity, akinesia, resting tremor and postural instability. It has recently been suggested that low frequency stimulation of the pedunculopontine nucleus (PPN) has a role in the therapy for Parkinsonism, particularly in gait disorder and postural instability. However, there is limited information about the mechanism of low frequency stimulation of the PPN on Parkinson's disease. The present study was to investigate the effect and mechanism of low frequency stimulation of the PPN on the firing rate of the ventrolateral thalamic nucleus (VL) in a rat model with unilateral 6-hydroxydopamine lesioning of the substantia nigra pars compacta. In vivo extracellular recording and microiontophoresis were adopted. The results showed that the firing rate of 60.71% VL neurons in normal rats and 59.57% VL neurons in 6-hydroxydopamine lesioned rats increased with low frequency stimulation of the PPN. Using microiontophoresis to VL neurons, we found the firing rate in VL neurons responded with either an increase or decrease in application of acetylcholine (ACh) in normal rats, whereas with a predominant decrease in M receptor antagonist atropine. Furthermore, the VL neurons were mainly inhibited by application of γ-aminobutyric acid (GABA) and excited by GABA(A) receptor antagonist bicuculline. Importantly, the VL neurons responding to ACh were also inhibited by application of GABA. We also found that the excitatory response of the VL neurons to the low frequency stimulation of the PPN was significantly reversed by microiontophoresis of atropine. These results demonstrate that cholinergic and GABAergic afferent nerve fibers may converge on the same VL neurons and they are involved in the effects of low frequency stimulation of the PPN, with ACh combining M(2) receptors on the presynaptic membrane of GABAergic afferents, which will inhibit the release of GABA in the VL and then improve the symptoms of Parkinson's disease.
Acetylcholine ; metabolism ; Action Potentials ; Animals ; Cholinergic Fibers ; physiology ; Electric Stimulation ; Male ; Oxidopamine ; Parkinson Disease, Secondary ; chemically induced ; physiopathology ; therapy ; Pedunculopontine Tegmental Nucleus ; physiology ; Rats ; Rats, Sprague-Dawley ; Ventral Thalamic Nuclei ; physiology

This study was aimed to investigate the protective effect of Liu Wei Dihuang (LWDH) against D-galactose (D-gal)-induced brain injury in rats and the existence of sex-dependent differences in LWDH protection. Sixty-four rats evenly composed of males and females were randomly assigned into 4 groups (n = 8): normal saline (NS) + NS (N + N), NS + LWDH (N + L), D-gal + NS (D + N) and D-gal + LWDH (D + L) groups. Rats in D + N and D + L groups received daily injection of D-gal (100 mg/kg, s.c.) for six weeks to establish the aging model, while rats in N + N and N + L groups were injected with the same volume of NS. From the third week, rats in N + L and D + L groups were orally administered with a decoction of LWDH for subsequent six weeks. Rats in N + N and D + N groups were orally administered just with the same volume of NS simultaneously. Morris water maze test was employed to evaluate the ability of learning and memory of the rats in all the groups. Acetylcholine (ACh) content, activities of choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) in visual cortex were assayed. Hematoxylin and eosin (HE) staining were used to observe the morphologic injury in hippocampus and visual cortex, and immunohistochemistry was performed to evaluate ChAT and AChE expression levels in the visual cortex. The results showed that the rats in D + N groups exhibited a longer escape latency to platform, lower swimming speed, less percent of target quadrant search time and platform crossings, compared with N + N groups, suggesting the establishment of aging model, while LWDH improved these indexes in D-gal-treated rats. Compared with D + N groups, LWDH increased ACh content and ChAT activity, and decreased AChE activity in visual cortex. Remarkable loss of neurons was found in hippocampus and visual cortex of aging rats, and the injury was significantly attenuated by LWDH. Immunohistochemistry showed D-gal-induced decreases of ChAT and AChE expressions were restored by LWDH. Furthermore, under the neural protection of LWDH, the improvement on platform crossings in male aging rats was better than that in female ones, while in ChAT expression and neuron density in visual cortex, female aging rats obtained more amelioration. These results suggest LWDH can markedly reverse the D-gal-induced cognitive impairments and neuronal damage in both hippocampus and visual cortex, which are achieved at least partly through restoring cholinergic system in central nervous system. Moreover, there is some sex difference in protective effects of LWDH against D-gal-induced impairment.
Animals ; Brain ; metabolism ; pathology ; Cholinergic Fibers ; drug effects ; pathology ; Cognition Disorders ; prevention & control ; Drugs, Chinese Herbal ; pharmacology ; Female ; Galactose ; toxicity ; Hippocampus ; metabolism ; pathology ; Male ; Neurons ; pathology ; Neuroprotective Agents ; pharmacology ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Visual Cortex ; metabolism ; pathology

OBJECTIVETo evaluate the effect of Gengnianchun Recipe (GNC) on learning memory function and its regulatory effect on hippocampal cholinergic system in ovariectomized rats.

METHODSFemale rats 10-12 months old were randomized into 5 groups, the sham-operation group, the model group treated with normal saline, the positive control group treated with Nilestriol, and the two GNC groups treated with high and low dose GNC respectively. A little fat around ovary was cut in the sham-operation group. The treatment lasted for 12 weeks after ovariectomy. Changes of learning memory function were tested by Morris water maze; serum level of estradiol (E2) was measured by chemical fluorescent method; hippocampal acetylcholinesterase (AChE) mRNA was determined with Real-time PCR; and the activities of acetylcholine (ACh), AChE and choline acetyltransferase (ChAT) in hippocampus were detected by immunohistochemistry respectively.

RESULTSTwelve weeks after ovariectomy, serum E2 and learning memory function markedly decreased in the ovariectomized rats (P < 0.01, P < 0.05). Nilestriol and high dose GNC showed an effect in improving the symptoms of learning memory functional deprivation and elevating the activities of hippocampal ACh, AChE and ChAT (all P < 0.05).

CONCLUSIONGNC can improve learning memory function of ovariectomized rats, and its mechanism might be realized by regulating the cholinergic system in hippocampus.

Acetylcholine ; metabolism ; Acetylcholinesterase ; genetics ; metabolism ; Animals ; Cholinergic Fibers ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Female ; Hippocampus ; drug effects ; metabolism ; Immunohistochemistry ; Maze Learning ; drug effects ; Memory ; drug effects ; Ovariectomy ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction

To determine the peripheral mechanisms involved in thermal sweating during the hot summers in July before acclimatization and after acclimatization in September, we evaluated the sweating response of healthy subjects (n=10) to acetylcholine (ACh), a primary neurotransmitter involved in peripheral sudomotor sensitivity. The quantitative sudomotor axon reflex test (QSART) measures sympathetic C fiber function after iontophoresed ACh evokes a measurable reliable sweat response. The QSART, at 2 mA for 5 min with 10% ACh, was applied to determine the directly activated (DIR) and axon reflex-mediated (AXR) sweating responses during ACh iontophoresis. The AXR sweat onset-time by the axon reflex was 1.50+/-0.32 min and 1.84+/-0.46 min before acclimatization in July and after acclimatization in September, respectively (p<0.01). The sweat volume of the AXR(1) [during 5 min 10% iontophoresis] by the axon reflex was 1.45+/-0.53 mg/cm2 and 0.98+/-0.24 mg/cm2 before acclimatization in July and after acclimatization in September, respectively (p<0.001). The sweat volume of the AXR(2) [during 5 min post-iontophoresis] by the axon reflex was 2.06+/-0.24 mg/cm2 and 1.39+/-0.32 mg/cm2 before and after acclimatization in July and September, respectively (p<0.001). The sweat volume of the DIR was 5.88+/-1.33 mg/cm2 and 4.98+/-0.94 mg/cm2 before and after acclimatization in July and September, respectively (p<0.01). These findings suggest that lower peripheral sudomotor responses of the ACh receptors are indicative of a blunted sympathetic nerve response to ACh during exposure to hot summer weather conditions.
Acclimatization ; Acetylcholine ; Axons ; Hot Temperature ; Iontophoresis ; Nerve Fibers, Unmyelinated ; Neurotransmitter Agents ; Receptors, Cholinergic ; Reflex ; Sweat ; Sweating ; Weather

OBJECTIVETo establish an artificial somatic-autonomic reflex arc in rats and observe the following distributive changes of neural fibers in the bladder.

METHODSAdult Sprague-Dawley rats were randomly divided into three groups: control group, spinal cord injury (SCI) group, and reinnervation group. DiI retrograde tracing was used to verify establishment of the model and to investigate the transport function of the regenerated efferent axons in the new reflex arc. Choline acetyltransferase (ChAT) in the DiI-labeled neurons was detected by immunohistochemistry. Distribution of neural fibers in the bladder was observed by acetylcholine esterase staining.

RESULTSDiI-labeled neurons distributed mainly in the left ventral horn from L3 to L5, and some of them were also ChAT-positive. The neural fibers in the bladder detrusor reduced remarkably in the SCI group compared with the control (P < 0.05). After establishment of the somatic-autonomic reflex arc in the reinnervation group, the number of ipsilateral fibers in the bladder increased markedly compared with the SCI group (P < 0.05), though still much less than that in the control (P < 0.05).

CONCLUSIONThe efferent branches of the somatic nerves may grow and replace the parasympathetic preganglionic axons through axonal regeneration. Acetylcholine is still the major neurotransmitter of the new reflex arc. The controllability of detrusor may be promoted when it is reinnervated by the pelvic ganglia efferent somatic motor fibers from the postganglionic axons.

Acetylcholinesterase ; biosynthesis ; Anastomosis, Surgical ; Animals ; Autonomic Fibers, Preganglionic ; physiology ; Cholinergic Fibers ; metabolism ; Immunohistochemistry ; Motor Neurons ; cytology ; metabolism ; Nerve Regeneration ; physiology ; Neural Pathways ; cytology ; metabolism ; Rats ; Rats, Sprague-Dawley ; Reflex ; physiology ; Spinal Cord Injuries ; physiopathology ; Spinal Nerve Roots ; surgery ; Urinary Bladder ; innervation ; physiology ; surgery ; Urinary Bladder, Neurogenic ; surgery

OBJECTIVETo investigate the general pattern of cholinergic nerve distribution and M(2) receptors in adult rat heart.

METHODSKarnovsky-Roots histochemical staining combining point counting method and immunochemical SABC method with image analysis were used to identify the cholinergic nerves and M(2) receptors, respectively, in adult rat heart.

RESULTSPositive staining of cholinergic nerves and M(2) receptors was found in all regions of the rat heart, and the point count of cholinergic nerves in the atria was 4.6 times as much as that in ventricles, and the area of immunoreactive substance for M(2) receptors two-fold higher in the atria than in the ventricles. The point counts of the cholinergic nerves in the medial-layer myocardium were fewer than that in subepicardial and endocardial tissues of the left ventricular free wall. However, M(2) receptors were comparable among the 3 layers of the left free ventricular wall.

CONCLUSIONCholinergic nerves and M(2) receptors are located in both rat atria and ventricles, but their density is much higher in the atria than in the ventricles. Transmural heterogeneity characterizes cholinergic nerve innervation in the left ventricular free wall without significant differences in M(2) receptor density.

Animals ; Cholinergic Fibers ; metabolism ; Female ; Heart ; innervation ; Heart Atria ; innervation ; metabolism ; Heart Ventricles ; innervation ; metabolism ; Immunohistochemistry ; Male ; Myocardium ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptor, Muscarinic M2 ; analysis

BACKGROUND: Cholinergic neurons in the forebrain of mamals are well elucidated, meanwhile, the distribution of these neurons in the brainstem is not conformed according to investigators. To identify cholinergic neurons in the brainstem of CD rat, the authors performed immunohistochemical staining with monoclonal antibody to choline acetyltransferase(ChAT), the enzyme responsible for the biosynthesis of acetylcholine. METHODS: In order to decrease nonspecific reactions of the secondary antibody, an anti-rat IgG without biotin was pretreated before the primary antibody treatment. Blockage of the protein transport was tried to decrease the secretion of ChAT from neurons with colchicine injection into the lateral ventricle of the rat 36, to 48 hours before cardiac perfusion of fixative agent. RESULTS: Choline acetyltransferase immunoreactive neurons were identified in all motor nuclei of cranial nerves with some other nuclei and nerve fibers in rat brainstem. They were all multipolar cells mostly composed of magnocellular neurons, however parvocellular ones were also observed with colchicine pretreatment. CONCLUSION: Distribution of the ChAT-immunoreactive nuclei in the brainstem of the CD rats were found in the area where motor functions are closely involved. This means that acetylcholine is the neurotransmitter of these area composing brainstem motor center.
Acetylcholine ; Animals ; Biotin ; Brain Stem* ; Choline O-Acetyltransferase ; Choline* ; Cholinergic Neurons ; Colchicine ; Cranial Nerves ; Humans ; Immunoglobulin G ; Immunohistochemistry ; Lateral Ventricles ; Nerve Fibers ; Neurons ; Neurotransmitter Agents ; Perfusion ; Prosencephalon ; Protein Transport ; Rats* ; Research Personnel