Suppressing SNAP-25 and reversing glial glutamate transporters relieves neuropathic pain in rats by ameliorating imbalanced neurotransmission / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Neuropathic pain results from a lesion or disease affecting the somatosensory system at either the peripheral or central level. The transmission of nociception within the central nervous system is subject to modulation by release and reuptake of neurotransmitters, which maintain a dynamic balance through the assembly and disassembly of the SNARE complex as well as a series of neurotransmitter transporters (inhibitory GABA transporters GAT and excitatory glutamate transporters GT). Neuronal hyper-excitability or defected inhibition involved in neuropathic pain is one of the outcomes caused by imbalanced neurotransmission. SNAP-25, which is one of the SNARE complexes, can modulate the release of neurotransmitters. Glia glutamate transporter (GLT) is one of the two glutamate transporters which account for most synaptic glutamate uptake in the CNS. The role of SNAP-25 and GLT as well as GAT is not clearly understood.</p><p><b>METHODS</b>We used the rat chronic constriction injury (CCI) model for research, and degraded SNAP-25 by a single intrathecal administration of BoNT/A. The mechanical (MWT) and thermal withdrawal latency (TWL) were tested. The level of SNAP-25, GLT, and GAT-1 were assayed using RT-PCR and Western blotting.</p><p><b>RESULTS</b>SNAP-25 was suppressed by a single intrathecal administration of 0.01U BoNT/A and the reduction of SNAP- 25 was correlated with the relief of nociceptive responses in CCI rats. MWT and TWL returned to normal from the 5th to 14th day (P < 0.05) after the administration. On the 14th day after surgery, compared to the sham group, the upregulation of SNAP-25 in CCI rats was reversed after BoNT/A treatment (P < 0.05). The decreased GLT was reversed after BoNT/A treatment but increased GAT-1 was not influenced by BoNT/A treatment.</p><p><b>CONCLUSIONS</b>SNAP-25 and GLT play important roles in the development of neuropathic pain, and the mechanism may involve the imbalance of neurotransmission after peripheral nerve injury. Intrathecal administration of BoNT/A reversed the upregulation of SNAP-25 and downregulation of GLT after CCI, but had no significant effect on the expression of GAT-1.</p>

Effect of intrathecal ketamine injection on protein kinase C expression in the spinal dorsal horn of rats with formalin-induced pain / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the expression of protein kinase C (PKC) in the spinal dorsal horn of rats with formalin-induced pain and the effect of intrathecal ketamine on PKC expression.</p><p><b>METHODS</b>Thirty-two SD rats were randomly divided into 4 equal groups, namely the control group, intrathecal saline group (NS), 50 µg ketamine group (K1) and 100 µg ketamine group (K2). The rats were anesthetized with 10% chloral hydrate, and a microspinal catheter was inserted intrathecally into the lumbar region. Five days later, the rats in groups, K1 and K2 were subjected to intrathecal administration of 50 and 100 µg ketamine (10 µl), respectively, followed by 10 µl saline, and those in NS group received 20 µl saline only. Thirty minutes later, 5% formalin (50 µl) was subcutaneously injected into the left hindpaw. The pain intensity score (PIS) was utilized to assess antinociceptive behavior within 1 h after formalin injection. Twenty-four hours later, the left hindpaw thickness was measured and the expression of PKC in the spinal dorsal horn in the L5 segment was assayed using immunohistochemistry.</p><p><b>RESULTS</b>Compared to group NS, groups K1 and K2 showed significantly decreased PIS (P<0.01) in the second phase of formalin-induced pain; 24 h later, the left hindpaw thickness of group NS increased obviously in comparison with that in the control group (P<0.01), whereas the thickness was significantly reduced in group K1 and K2 as compared to that in group NS (P<0.05). The number of immunoreactive cells and the immunohistochemical score of PKC in the spinal dorsal horn were significantly higher in group NS than in group C (P<0.01), but significantly lower in groups K1 and K2 than in group NS (P<0.05).</p><p><b>CONCLUSION</b>Intrathecal ketamine produces obvious antinociception against formalin-induced pain in rats and inhibits the enhanced PKC expression in the spinal dorsal horn in response to formalin-induced pain, suggesting the important role of PKC in nociceptive signal transmission and modulation in the spinal cord.</p>

Three subanaesthetic dose ketamines mixed with butorphanol in the postoperative continuous intravenous analgesia / 中南大学学报(医学版)

OBJECTIVE@#To determine an optimal clinical dose of ketamine after comparing the efficacy and security of 3 low dose ketamines mixed with butorphanol in the postoperative continuous intravenous analgesia.@*METHODS@#Eighty ASA (American Society of Anesthesiologists) I-II patients scheduled for elective gynecological surgery under general anesthesia were divided randomly into 4 groups (n=20): Group B received butorphanol 3 microg/(kg x h);Group BK1 received butorphanol 2 microg/(kg x h) mixed with ketamine 60 microg/(kg x h); Group BK2 received butorphanol 2 microg/(kg x h) mixed with ketamine 90 microg/(kg.h); and Group BK3 received butorphanol 2 microg/(kg x h) mixed with ketamine 120 microg/(kg x h). Continuous intravenous infusion pump was used when the patients had obvious pain (visual analgesia scale of five), and the bolus infusion (4 mL) was given before the operation, and continuous infusion at 2 mL/h. In the postoperative period, pain was assessed using visual analogue scale (VAS) at 2,6,12,24, and 48 h.At the same time, Ramsay scores and adverse effects were recorded.@*RESULTS@#There was no significant difference in the adverse effects and the postoperative mean arterial pressure, heart rate, respiratory rate values, and pulse oxygen among the 4 groups. Postoperative VAS values in Group BK3 was the lowest, followed by Group BK2. There was no significant difference between Group BK1 and Group B. The incidence of somnolence in Group B was higher than that in Group BK1, BK2 and BK3(P<0.05).@*CONCLUSION@#Ketamine in subanaesthetic dose added to butorphanol for postoperative continuous intravenous infusion has a better postoperative analgesic effect and sedation. It can effectively spare butorphanol consumption without increasing adverse effects. The optimal combined dose is 90-120 microg/(kg x h).

Effect of intrathecal pumping tramadol on the immune function in rats with formalin pain / 中南大学学报(医学版)

OBJECTIVE@#To evaluate the effect of intrathecal pumping tramadol on cell-mediated immunity in rats with formalin inflammatory pain.@*METHODS@#Thirty-two Sprague-Dawley adult male rats weighting 250 approximately 300 g were randomly divided into 4 groups (n=8 in each group):Saline group (NS) and 3 tramadol groups (T1,T2,and T3). The rats were anesthetized with intraperitoneal chloral hydrate (300 approximately 350)mg/kg. Microspinal catheter was inserted into the subarachnoid space at the lumber region according to modified Yaksh techniques. In the tramadol groups,after 5 days tramadol was continuously infused through the spinal catheter at 50 (T1),25 (T2), and 12.5 microg/h (T3) for 7 days. In the NS group normal saline was continuously infused instead of tramadol. On Day 7 formalin (5%, 50 microL) was injected into the plantar surface of the left hindpaw. The number of flinches, lickings and total time of licking was recorded for 60 min.Pain intensity scoring(PIS)(0 approximately 3;0= no pain, 3=severe pain) was used to assess the antinociceptive effect of intrathecal tramadol. The rats were killed after the evaluation of pain intensity. Body weight and spleen weight were measured and spleen index (spleen weight/body weight) was calculated. T-lymphocyte function was evaluated based on Concanavalin-A(ConA) induced splenocyte proliferation. A modified lactic acid dehydrogenase(LDH) release assay was done to assess the NK cell activity. Phenotypic expressions of cell surface markers of T lymphocyte subsets (CD3+, CD3+ CD4+, CD3+ CD8+, and CD4+/ CD8+) and NK cell(CD161+) in the spleen were analyzed by flow cytometry.@*RESULTS@#The PIS scores were significantly lower in the T1,T2,and T3 groups than those in the NS group. The spleen index and splenocyte proliferation induced by ConA were significantly suppressed in the T1 group,and the phenotypes of T lymphocyte subsets were significantly changed,but no significant difference was found in the T2 and T3 groups compared with the NS group. There were no differences in NK cell activity in the 3 tramadol groups from the control group.@*CONCLUSION@#Intrathecal pumping tramadol has significantly antinociceptive effect. Intrathecal pumping higher dosage tramadol (50microg/h) suppresses T lymphocyte proliferation and alteres T lymphocyte subset phenotype but does not affect NK cell activity. General analgesic dosage tramadol (25 and 12.5 microg/h) has no effect on the immune function.

Periaqueductal gray administration of HSV-I amplicon vector-mediated HPPE gene therapy of nocicepion in rats with formalin-induced pain / 中南大学学报(医学版)

OBJECTIVE@#To investigate the antinociceptive effect of periaqueductal gray (PAG) administration of herpes simplex virus type-1(HSV-I) amplicon vector-mediated human preproenkephalin gene (HPPE).@*METHODS@#Sprague-Dawley rats weighting 260 to approximately 320 g were randomly divided into pHSVIRES-HPPE-LacZ (SHPZ) group, pHSVIRES-LacZ (SHZ) group, and saline (NS) group which included 3 d,1 week,2 week,3 week,4 week,5 week, and 6 week groups (n=51). The rats were anesthetized with intraperitoneal chloral hydrate (300 to approximately 350) mg/kg. Rats were PAG delivered with recombinant HSV-I amplicon vector SHPZ, SHZ or NS. One week after PAG administration 9 rats in each group were sacrificed and lumber segment of the spinal cord was removed for determination of expression of LacZ by X-gal staining and HPPE mRNA expression by reverse transcription-polymerase chain reaction and L-enkephalin content by radioimmunoassay in PAG. Formalin 50 microL (5%) was injected into the left hindpaw, and pain intensity scoring (PIS) was used to assess the antinociceptive effect.@*RESULTS@#After in vivo transferring, neurocyte demonstrated strong positive signals with X-gal immunohistochemical staining. The expression of HPPE mRNA was detected in PAG after administration of SHPZ. PAG delivery of SHPZ showed antinociceptive effect on formalin-induced pain for 6 weeks compared with SHZ group.@*CONCLUSION@#This amplicon virus can transfer HPPE into rat PAG neural cells and make it express efficiently. PAG administration of SHPZ can produce significant analgesic effect on formalin-induced pain in rats for 5 weeks.

Intrathecal injection of ketamine and clonidine for chronic neuropathic pain model in rats / 中南大学学报(医学版)

OBJECTIVE@#To observe the effect of intrathecal injection of ketamine and clonidine for chronic constriction injury (CCI) in rats.@*METHODS@#Thirty-two SD male rats weighing 220-280 g were anesthetized with intraperitoneal chloral hydrate 300 mg/kg. A catheter was implanted in the subarachnoid space at the lumbal region and CCI rat models were made successfully. On the 4th day after the surgery, the rats were randomly divided into 4 group: a control group,injecting 0.9%NS 20 microL intrathecally; a ketamine group, injecting ketamine 1 mg/kg(20 microL) intrathecally; a clonidine group (CL), injecting clonidine 20 microg/kg (20 microL) intrathecally; a combined ketamine and clonidine group, injecting ketamine 0.5mg/kg and clonidine 10 g/kg (20 microL) intrathecally, once a day for 1 week. BME-410A Plantar Analgesia Tester was used to measured pain threshold before the administration and 30 min after the administration. The rats were killed after the test was finished. And then we detected the nitric oxide synthase (NOS) activity and the NO production in the spinal cord.@*RESULTS@#The combined injection of ketamine (0.5mg/kg)and clonidine(10 g/kg) produced significantly more potent analgesia than the injection of ketamine (1 mg/ kg) or clonidine (20 microg/ kg)alone. The NOS activity and the production of NO in the combined injection group were significantly lower than those of the single injection group (P<0.05). The weight of rats post-administration increased obviously in the 4 groups (P<0.05).@*CONCLUSION@#The combined injection of ketamine and clonidine can produce synergistic ab-irritation without obvious side effects.

Intrathecal administration of HSV-I amplicon vector-mediated HPPE gene therapy of nocicepion in rats with formalin pain / 中南大学学报(医学版)

OBJECTIVE@#To investigate the antinociceptive effect of intrathecal administration of HSV-I amplicon vector-mediated HPPE.@*METHODS@#Sprague Dawley rats (290+/-30) g were randomly divided into pHSVIRES-HPPE-LacZ (SHPZ) group, pHSVIRES-LacZ (SHZ) group, and saline group (NS), and 3 d, 1 week, 2 weeks, 3 weeks, 4 weeks, and 5 weeks group,which were anesthetized with 10% chlroral hydrate 300- 350 mg/kg. A microspinal catheter was inserted into the lumbar subarachnoid space. Rats were intrathecally delivered with recombinant HSV-I amplicon vector SHPZ, SHZ or NS. The HPPE expression was detected by reverse transcription-polymerase chain reaction (RT-PCR) and radioimmune assay. Formalin 50 microL (5%) was injected into the left hindpaw, pain intensity scoring (PIS) was used to assess the antinociceptive effect.@*RESULTS@#After in vivo transferring,neurocyte demonstrated strong positive signals with X-gal immunohistochemical staining. RT-PCR and L-enkephalin radioimmune assay found that the neural cells transferred foreign gene (HPPE) had effective expression. Intrathecal delivery of SHPZ showed antinociceptive effects on formalin induced pain for 5 weeks compared with SHZ.@*CONCLUSION@#This amplicon virus can transfer HPPE into rat central nerve system neural cells and express efficiently, suggesting SHPZ is satisfactory treatment for gene therapy for chronic pain. Intrathecal delivery SHPZ demonstrated antinociceptive effects on formalin induced pain.

Effect of intrathecal katemine on the expression of nNOS in spinal dorsal horn in rats with formalin pain / 中南大学学报(医学版)

OBJECTIVE@#To investigate the expression of neuronal nitric oxide synthase (nNOS) in spinal dorsal horn and the effect of intrathecal katemine on the expression of nNOS in the rats with formalin-induced pain.@*METHODS@#Thirty-two Sprague-Dawley rats were randomly divided into 4 groups (n=8 in each group): a control group (C), an intrathecal saline group (NS), a katemine 50 microg group (K1), and a katemine 100 microg group (K2). The rats that were anesthetized with 10% chlroral hydrate 300 - 350 mg/kg by abdominal injection were intrathecally inserted a microspinal catheter into the lumbar region according to the method of modified Yaksh. After 5 days ,the rats in Group NS, K1 and K2 were intrathecal 20 microL saline or 10 microL katemine (50, 100 microg) followed by 10 microL saline, respectively. Thirty minutes later, 5% formalin 50 microL was subcutaneously injected into the left hindpaw. Pain intensity scoring (PIS) was used to assess antinociceptive behavior within 1 h after the formalin injection. The expression of nNOS in the spinal dorsal horn of L5 segment was assayed using immunohistochemistry 24 h later.@*RESULTS@#Compared with Group NS, PIS of Group K1 and K2 was decreased obviously (P<0.01) in the second phase of formalin pain. The number of immunoreactive soma and immunohistochemistic dying grade of nNOS in the spinal dorsal horn of L5 segment was higher in Group NS than that in Group C (P<0.01), but Group K1 and K2 were lower than Group NS (P<0.01).@*CONCLUSION@#There was significant antinociception of intrathecal katemine in rats with formalin pain. Intrathecal katemine significantly inhibited the increase of nNOS expression in the spinal dorsal horn of formalin-induced pain, suggesting nNOS plays an important role in nociceptive transmission and modulation of the spinal cord.

Spinal anesthesia with low dose sufentanil-bupivacaine in transurethral resection of the prostate / 中南大学学报(医学版)

OBJECTIVE@#To explore the clinical efficacy of intrathecally administered low dose sufentanil-bupivacaine in transurethral resection of the prostate (TURP). METHODS. Ninety patient (ASA I - III) undergoing TURP were randomly divided into 3 groups (n = 30); Group A, B and C. Group A received 7.5 mg bupivacaine + 5 microg sufentanil + 10% glucose; Group B received 7.5 mg bupivacaine + 7.5 microg sufentanil + 10% glucose; Group C received 15 mg bupivacaine + 10% glucose. The volume was 3 mL in every group. SP, DP, HR, SpO2, the degree of motor and sensory blockade and the side effect were observed.@*RESULTS@#SP/DP was significantly decreased in Group C than that in Group and Group B (p<0.05), HR and SpO2 in group B were decreased to different degrees 15 min after the injection (p<0.05). The complete recovery time of motor nerve blockade and the regression time of sensory blockade were obviously prolonged in Group C (p<0.05). There were no significant differences in analgesic effect among the three groups during the operation, but the incidence of pruritus was higher in both group A and Group B than that in Group C during the first 24 hours after the injection.@*CONCLUSION@#Spinal anesthesia with low dose sufentanil-bupivacaine possesses relatively steady hemodynamics. The blockade degree of motor and sensory blockade in this spinal anesthesia is lower than that in standard spinal bupivacaine in TURP.

Clinical study on bispectral index in monitoring the depth of desflurane anesthesia in the elderly / 中南大学学报(医学版)

OBJECTIVE@#To evaluate the values of bispectral index (BIS) in mornitoring the depth of desflurane anesthesia in the elderly by observing the changes of BIS at various end-tidal desflurane concentrations.@*METHODS@#BIS was used to monitor the depth of desflurane anesthesia in the elderly without surgery stimulation. Forty ASA physical status I approximately III patients undergoing general anesthesia were divided into 2 groups with 20 in each group:Group Elderly ( > or =65 years) and Group Youth ( 18 approximately 55 years). Anesthesia was induced with propofol 2. 0 mg/kg and vecuronium 0. 1 mg/kg including the endotracheal topical anesthesia with 2% lidocaine. After the desflurane in oxygen, each concentration of desflurane was maintained for 20 minutes. The changes of mean arterial blood pressure (MAP), heart rate (HR) and BIS were recorded simultaneously. The timepoints setting for observation were: preanesthesia, 2 minutes after the anesthesia, endotracheal intubation, 2 minutes after the intubation, and end-tidal concentration of desflurane at 0. 6 MAC, 1.0 MAC and 1. 3 MAC.@*RESULTS@#During anesthesia of desflurane, MAP and HR did not change significantly in the 2 groups with increasing end-tidal desflurane concentration from 0. 6 MAC to 1. 3 MAC (P > 0.05). BIS decreased significantly than that at preanesthesia in the 2 groups during anesthesia of desflurane ( P 0. 05 ). In general, BIS highly correlated with the end-tidal desflurane concentration. The coefficient of product-moment correlation (r) between BIS and the end-tidal desflurane concentration was -0. 996 and -0. 946 in Group Elderly and Youth (P < 0. 05 ).@*CONCLUSION@#BIS highly correlates with the end-tidal desflurane concentration which is used to evaluate the depth of desflurane anesthesia in the elderly and youth. There is different depth of anesthesia by BIS in the elderly or youth at the same end-tidal desflurane concentration.

Behaviorial observation of ropivacaine in continuous spinal anesthesia and Ca2+ content of spinal cord in rats / 中南大学学报(医学版)

OBJECTIVE@#To observe the behaviour and Ca2+ content of spinal cord in rats after continuous spinal anesthesia administration of different densities and doses of ropivacaine in SD rats.@*METHODS@#Twenty-four male SD rats weighing 220 approximately 280 g were anesthetized. A polyurethane microspinal catheter was inserted into the lumbar subarachnoid space 8 cm according to the method of Yaksh's. The animals were randomly divided into 4 groups of 6 each: in group N the rats were given normal saline 40 microl intrathecally every one and half hours for 3 times; in group R1, 0.5% ropivacaine was given; in group R2 0.75% ropivacaine and in group R3 1% ropivacaine was given. The activity of rats was observed. After 6 hours rats were perfused with 4% formamint through the ascending aorta. The rats were sacrificed and L1 approximately 2 segment of spinal cord was immediately removed for Ca2+ content examination.@*RESULTS@#A total hind limbs paralysis was seen at 30 seconds and intramuscular strain gradually came back from 30 to 90 minutes after the intrathecal administration of ropivacaine in all rats. The recovery of motor black was remarkably different in group R1, R2, and R3 (P < 0.05). The Ca2+ content of spinal cord was significantly higher in group R3 than that in group R1 and R2 (P < 0.05 ).@*CONCLUSION@#There is no significant change of motor black time and it is related to drug dose for 0.5% , 0.75% and 1% ropivacaine in continuous spinal anesthesia. 1% ropivacaine may increase Ca2+ content in spinal cord.

Effect of intrathecal pumping morphine on immunological function in rats with formalin pain / 中南大学学报(医学版)

OBJECTIVE@#To evaluate the immunological function in rats with formalin inflammatory pain through intrathecal pumping different dosages of morphine.@*METHODS@#Thirty-two Sprague-Dawley rats were randomly divided into 4 groups (n = 8 in each group): saline group (NS) and morphine group included M1 group (10 microg/h) , M2 group (5 microg/h), and M3 group (2.5 microg/h). Chronic intrathecal catheterization was performed under anesthesia with 10% chloral hydrate (300-350) mg/kg according to M2 group (5 microg/h) and M3 group (2. 5 microg/h). Chronic intrathecal catheterization was modified Yaksh's. After 7 days, pain intensity scoring (PIS) was utilized to assess antinociceptive effect of morphine. And spleens were aseptically removed to obtain splenic cells. T lymphocyte function was evaluated based on Concanavalin-A induced splenocyte proliferation. A modified lactic acid dehydrogenase release assay was used to assess NK cell activity. Phenotypic expression of cell surface markers of T lymphocyte subsets (CD3+, CD3+ CD4+, CD3+ CD8+, and CD4+ / CD8+ ) and NK cell ( CD161+) in the spleen were analyzed by flow cytometry.@*RESULTS@#Compared with the NS group, PIS of morphine group decreased obviously (P < 0.01) and was dose-dependent in the early and late phase of formalin pain, but there were no significant differences among morphine groups. Spleen index, splenocyte proliferation and NK cell activity were significantly suppressed by intrathecal pumping morphine. Phenotypic expression of T lymphocyte subsets and NK cell assessed by flow cytometry were different from the control group in all morphine groups.@*CONCLUSION@#There was significant antinociception of intrathecal pumping morphine. After intrathecal pumping different dosages of morphine (10 microg/h,5 microg/h, and 2.5 microg/h), the function of cellular immunity was suppressed and was dose-dependent.