To identify the role of spinal cyclooxygenase (COX)-1 in the development and maintenance of postoperative pain, we examined the changes of COX-1 protein expression in lumbar spinal cord by immunohistochemistry and Western blot technique in rat plantar incision model at different time points (pre-incision or 2 h, 4 h, 6 h,12 h, 1 d, 2 d, 3 d, 5 d and 7 d after incision). We also studied the anti-allodynic effects of the COX inhibitors by intrathecal administration of non-selective COX inhibitors (ketorolac), selective COX-1 (SC-560) or COX-2 inhibitor (NS-398) immediately or 2 h, 24 h after incision. The mechanical allodynia was evaluated by using paw withdrawal threshold (PWT) response to mechanical stimulation on pre-incision, 2 h, 1 d, 2 d, 3 d, 5 d and 7 d after incision or 30 min after drug treatment. The result showed that COX-1 immunoreactive cells mainly focused in the superficial laminae of lumbar spinal dorsal horn and expression of spinal COX-1 protein increased after incision, peaked at 4 h (P＜0.01) and lasted for 12 h. Postoperative treatment with both SC-560 and ketorolac significantly alleviating the mechanical allodynia induced by skin incision, but NS-398 had no such effect. This study demonstrates that spinal COX-1 involves in the development and maintenance of postoperative hypersensitivity and intrathecal COX-1 inhibitor has anti-allodynic effect on incision pain in the rat.

Whole-cell patch clamp technique was performed on acutely isolated rat dorsal root ganglion (DRG) neurons to investigate the modulatory effect of caffeine on γ-aminobutyric acid (GABA)-activated currents (IGABA). The results showed that the majority of the neurons examined (97.4%, 113/116) were sensitive to GABA. 1-1000 μmol/L GABA activated a concentration-dependent inward current which manifested obvious desensitization. After the neurons were treated with caffeine (0.01-100 μmol/L) prior to the application of GABA (100 μmol/L) for 30 s, GABA-activated membrane currents were obviously inhibited. Caffeine shifted the GABA dose-response curve downward and decreased the maximum response to 57% without changing Kd value. These results indicate that the inhibitory effect is non-competitive. Theophylline showed a similar and stronger inhibitory effect on IGABA. The pretreatment with caffeine (10 μmol/L) inhibited IGABA, which was potentized by diazepam (1 μmol/L). Intracellular application of H-8 almost completely abolished the inhibitory effect of caffeine on IGABA. The present results suggest that caffeine may be able to antagonize the effect of presynaptic inhibition of GABA in primary afferent.

Effects of urotensin II (UII) on paraventricular nucleus (PVN) neurons of hypothalamus from brain slices of rats were examined by using extracellular recording technique. The results are as follows: (1) In response to application of UII (0.3, 3.0, 30.0, 300.0 nmol/L, n=39) into the perfusate for 2 min, the spontaneous discharge rates (SDR) of 32/39 (82.05% ) neurons were significantly decreased in a dose-dependent manner; (2) Pretreatment with bicuculline (BIC, 100 μmol/L), a specific GABAA receptor antagonist, led to a marked increase in SDR of 5/7 ( 71.43% ) neurons in an epileptiform pattern. The increased discharges were not significantly changed after UII ( 30.0 nmol/L ) was applied into the perfusate for 2 min; (3) Pretreatment with picrotoxin ( PIC, 50 μmol/L ), a selective blocker of Cl- channel, led to an increase in the SDR of all 12/12 (100%) neurons. The increased discharges were not influenced by the applied UII (30.0 nmol/L) for 2 min in 11/12 (91.67%) neurons; (4) Application of nitric oxide synthase (NOS) inhibitor NG-nitro-L-arginine methyl ester (L-NAME, 50 μmol/L ) into the perfusate could significantly augment the SDR of 11/12 ( 91.67% ) neurons , while UII ( 30.0 nmol/L ) applied into the perfusate for 2 min led the augmented SDR of all (12/12, 100%) neurons decrease. The results suggest that UII decreases the excitability of PVN neurons of hypothalamus by potentiating GABAA receptor-mediated Cl- current.

Previous experiments has shown that Ginsenoside Rb1 (GRb1), which is one of the most important active ingredients in ginseng (Panax ginseng C.A. Meyer), reduced infarct and neurologic deficit followed by the transient cerebral ischemia in rats. The mechanism of this neuroprotective function is unclear. In this study, we tested whether the neuroprotective effect of GRb1 is achieved through preventing the neuronal apoptosis and modulating expression of neuronal apoptosis inhibitory protein (NAIP). Focal cerebral ischemia was induced by the middle cerebral artery occlusion (MCAO) in Wistar rats. GRb1 (40 mg/kg, i.p.) was administered immediately after the onset of reperfusion. The rats with neurological deficits were randomly divided into 2 groups: the ischemia and the GRb1 group. Each group was again divided into subgroups according to the various reperfusion time (3 h, 12 h, 1, 2, 3, 5, 10 days, n=4 per time point). Apoptotic cells were analyzed using TUNEL. Immunohistochemical method was used to assess expression of NAIP. This results showed that the number of apoptotic cells elevated at 3 h of reperfusion, and peaked at 24 h, then declined, but the number of apoptotic cells at 10 d after ischemia was significantly more than those of control groups (P＜0.01). Compared with ischemia group, the apoptotic cells decreased at all subgroups of GRb1; however, the significant differences were only found from 12 h to 3 d of reperfusion. In normal and sham groups, NAIP weak immunostaining was diffusely present in the neurons of parenchyma. The number of NAIP-positive cells started to increase in ischemic regions at 3 h after ischemia, peaked at 12 h and declined up to 5 d of reperfusion. At 5 d after ischemia, the number of NAIP-positive cells was less than that of control group (P＜0.05). A few astrocytes strongly expressed NAIP in the ischemic area. In the GRb1 group, the number of NAIP-positive cells from 12 h to 10 d after ischemia was evidently higher than in the ischemia group. Thus, these results suggest that GRb1 has potential ability to prevent apoptosis, the mechanism of which is related to induce expression of NAIP.

To investigate the relationship of the rotation stimulation with motion sickness, the expression of Fos protein in the whole brain of the rat stimulated by complex double rotation on two axes was observed in the present study. The rats were randomly divided into four groups: normal contral group; double-axes rotation stimulation group; the bilateral labyrinthectomy group; group of two-axes rotation stimulation after the bilateral labyrinthectomy. Immunohistochemical staining method was used to detect the expression of Fos protein in different regions of whole brain of the rat. The present results showed that: (1) No Fos-like immunoreactivity was detected in the brain of the rats in control group and the bilateral labyrinthectomy group; (2) In the double-axes rotation stimulation group, the Fos-like immunoreactive neurons were observed in many regions of the brain and brainstem of the rats following complex double-axes rotation stimulation, and the Fos-immunoreactivities were expressed in the nucleus. These Fos-immunopositive neurons were intensively distributed in different subnuclei of the vestibular nucleus complex (including medial, superior and spinal nuclei), nucleus of the solitary tract, locus coeruleus, medial and lateral parabrachial nucleus of the brainstem, paraventricular nucleus of the diencephalons and the amygdala of the limbic system; (3) The expression of Fos protein can be scarcely detected around forementioned regions in brains of the rats following complex two-axes rotation stimulation after the bilateral labyrinthectomy. The present results suggest that the double-axes rotation stimulation can activate effectively the vestibular neurons and many neurons of other region of the brain and brainstem are further activated through direct or indirect connections with vestibular nuclei after complex double rotation stimulation. These activated neurons may be related to the complex mechanism of the motion sickness.

We examined the effects of intracerebroventricular ( i. c. v) administration of adrenomedullin (ADM) on catecholaminergic neurons and the expression of c-fos gene in rat brain nuclei involved in cardiovascular regulation using double immunohistochemical method for Fos and tyrosine hydroxylase (TH). The results showed that: ( 1 ) Following icy administration of ADM (3 nmol/kg) , double-labeled neurons for Fos and TH were significantly increased in the area postrema ( AP), the nucleus of the solitary tract ( NTS), the nucleus paragigantocelluaris laterialis (PGL) and the locus coeruleus (LC). (2) Pretreatment with calcitonin gene-related peptide receptor antagonis CGRP8-37 (30 nmol/kg) significantly reduced the action of ADM (3 nmol/kg) in the brain. The present study suggested that ADM might activate the neurons of the brain nuclei involved in cardiovascular regulation, and supported the hypothesis that the central action of ADM were induced by activating the catecholaminergic neurons of brainstem nuclei involved in cardiovascular regulation, CGRP receptor might mediate the effects of ADM.

To investigate the expression change of BDNF in lamina II of spinal cord from partial deafferented cats, L6 segmentsof spinal cord from 20 adult male cats (5 normal cats, 15 unilateral L6 spared roots cats allowed to survive 3 d, 6 d and 12 d re-spectively) were stained with immunohistochemical technique. The results showed: BDNF positive products were mainly dis-tributed in nerve terminals, varicosities and few neurons of spinal cord lamina II in normal cat. After operation, the density ofpositive nerve terminals and varicosities began to decrease on the third day, reached the lowest level on the 6th day and recoveredto normal level on the 12th day on operated side. But the number of BDNF neurons showed no obvious change. The authors sug-gest that the decreased density of BDNF positive products in lamina II on the 3rd and 6th day was related with the degenerationof the nerve fibers and varicosities after section of the adjacent dorsal roots. On the 12th day, the remaining L6 dorsal roots un-derwent collateral sprouting compensatoryly and reestablished functional connection with target neurons. Therefore, BDNF maybe involved in the normal physiological function and the plasticity of spinal cord after damage.

To understand the relationship of neurotransmitter between the striatum and limbic system such as amygdaloid nucle-us and bed nucleus of the stria terminalis. 30 male Sprague Dawley rats were used. Immunohistochemical ABC method was per-formed to detect the expression of substance P (SP), calcitonin gene-related peptide (CGRP), leucine-enkephalin (L-enk),cholecyctokinin (CCK) and neuronal nitric oxide synthase (NOS) on seetions of the brain. Some transmitters including substanceP, calcitonin gene-related peptide and cholecyctokinin were mainly distributed at the marginal division of the striatum. Theleucine-enkephalin was mainly distributed at the globus pallidus and was secondly distributed at the marginal division of the stria-tun. The neuronal nitric oxide synthase was mainly distributed at caudate putamen and the marginal division. All these transmit-ters were not only distributed at amygdaloid nucleus and bed nucleus of the stria terminalis, but also had fibers connection amongthe amygdaloid nucleus, marginal division and bed nucleus of the stria terminalis. CONCLUSION: There were special fibersconnection between the marginal division and other basal ganglia nucleus or the limbic system. The marginal division may beplayed some important functions of basal ganglia and limbic system.

In order to investigate the chemical and morphornetrical properties of the neuronal precursor cells derived from thesubventricular zone(SVZ) of the postnatal rat forebrain in vitro. The cell-type specific antibodies were used for the immunocy-tochemical staining ,and the morphometric parameters which were the mean soma diameter and the ellipticity index (i. e. , thesmallest soma diameter divided by the largest soma diameter) of every SVZ-derived cell were measured for identifying the pheno-types of the SVZ cells in vitro. The experiment animals were SD rats (weights: 100～ 150 g), the SVZ cells derived from thepostnatal rats were cultured on poly-D-lysine-coated 24-well glass chamber slides in the Neurobasal Medium supplemented withB27 in 5% CO2 at 37 C. The following results were obtained.. At 1 day in vitro, almost all SVZ cells (〉90%) from the postna-tal rat forebrain expressed Tujl, an antibody that recognizes neuron-specific tubulin. Likewise, the preponderance of the SVZcells expressed the polysialylated neural cell adhesion molecule (PSA-N-CAM) ; The majority of the SVZ Tujl-positive cells cul-tured were the cells that had oval-shaped bodies with two short, unbranched processes protruded from every two poles, theirmean soma diameter were 8.42±1.03μm and their ellipticity index were 0.57±0.12. Meanwhile, there were approximately20% of the SVZ cells in culture that were sphere-shaped cells with mean soma diameter 7.20±l.04 μm , and it might be observed that these cells connected with one another. As the time in culture went on, these sphere-shaped SVZ-derived cells alsotransformed to oval-shaped ones as described above, but it could be observed that the cells were still connected in the processesof them. By 3 and 5 days in culture, the SVZ cells had larger cell somas (average diameter 9. 07±1.07 μm), and often consider-ably longer processes but still with few branches. Immunocytochemical staining revealed that the majority of the SVZ cells in cul-ture remained Tujl-positive, PSA-N-CAM-positive. By 7 days in culture, the Tujl-positive cells in culture showed remarkablemorphological changes, and possessed typical neuronal phenotypes, which had more larger cell somas (average diameter 12.8 ±1.13 μm), and had more longer, branched processes. Our results showed that the SVZ in the postnatal SD rats contained theneuronal precursor cells which were PSA-N-CAM-positive and could differentiate into new neurons in vitro.

After injecting retrograde tracer fiuoro-gold (FG) into the parabrachial region(PB), caudal ventrolateral medulla(CVLM) and the fourth segment of cervical spinal cord (C4), respectively, neurons in laminae I ～ Ⅱ of the medullary dorsalhorn projecting to the above mentioned brain areas were observed. PB received projections from bilateral laminae I and Ⅱ withan ipsilateral dominance; CVLM and C4 received projections from ipsilateral laminae I and Ⅱ. Neurons projecting to C4 werevery sparsely distributed in laminae I and Ⅱ of the medullary dorsal horn. The projecting neurons in outer part of lamina Ⅱwere more than those in inner part of lamina Ⅱ . Combined with immunofluorescence histochemistry for calbindin-D28k(CB) andparvalbumin(PV), it was demonstrated that a part of neurons projecting to PB or CVLM showed CB-like immunoreactivity, butnone of them exhibited PV-like immunoreactivity. There were only a few neurons in lamina Ⅱ projecting to C4 and they exhibitedneither CB- nor PV-like immunoreactivity. The present study provides further evidence for the existence of projecting neurons inlamina Ⅱ and suggests that immunostaining against CB and PV may distinguish two neuronal subpopulations in lamina Ⅱ .