Re-exposure to morphine-associated context facilitated long-term potentiation in the vSUB-NAc glutamatergic pathway via GluN2B-containing receptor activation.

The glutamatergic projection from the ventral subiculum of the hippocampus (vSUB) to the nucleus accumbens (NAc) shell has been reported to play a key role in drug-related behavior. The GluN2B subunit of N-methyl-D-aspartate receptors (NMDARs) in the NAc can be selectively elevated after the retrieval of drug-conditioned memory. However, whether the increased GluN2B-containing NMDARs (GluN2B-NMDARs) are able to alter the synaptic plasticity of the vSUB-NAc glutamatergic pathway remains unclear. Here, we found that the long-term potentiation (LTP) in the vSUB-NAc pathway was facilitated and the GluN2B subunit protein level was elevated in synaptoneurosomes of the NAc shell, but not in the core, following morphine-induced conditioned place preference (CPP) expression in rats. The facilitated LTP was prevented by the GluN2B-NMDAR antagonist RO25-6981. Also, a neurochemical disconnection following microinjection of RO25-6981 into the NAc shell, plus microinfusion of GABA agonist baclofen and muscimol into the contralateral vSUB prevented the expression of morphine-induced CPP. These findings suggest that the retrieval of drug-associated memory potentiated synaptic plasticity in the vSUB-NAc pathway, which was dependent on GluN2B-NMDAR activation in the NAc shell. These findings provide a new explanation for the mechanisms that underlie the morphine-associated-context memory. The GluN2B-NMDARs may be regarded as a potential target for erasing morphine-related memory.

Transcutaneous electrical acupoint stimulation for the treatment of withdrawal syndrome in heroin addicts.

OBJECTIVE: To assess the therapeutic effect of transcutaneous electric acupoint stimulation (TEAS) for the treatment of withdrawal syndrome in heroin addicts. METHODS: A total of 63 male heroin addicts with withdrawal score higher than 20 were recruited in the Detoxification Center of Zhongshan city, Guangdong province, China. They were randomly distributed into two groups: TEAS group (n = 31) received TEAS by using a Han's acupoint nerve stimulator (HANS) model 200A with two output channels, 2-3 sessions per day, 30 minutes per session for 10 consecutive days. Electrical stimulation of alternating frequencies of 2- and 100-Hz with 3 second each, and with intensity of 10-15 mA was applied on Hegu (LI-4) and Laogong (PC-8) points on one hand, and Neiguan (PC-6) and Waiguan (SJ-5) points on the other forearm via electroconductive skin pads of 4 cm × 4 cm in size. The control group (n = 32) was treated with similar procedure except that the leads of the output of the stimulator was disconnected. Assessments of the severity of the withdrawal syndrome were conducted one day before and on each day during the whole treatment period of 10 days. Buprenorphin of 1 mg per day sublingually was provided to all subjects in the first two days, and then to those with withdrawal score over 20 in the following days. RESULTS: The TEAS treatment dramatically alleviated the withdrawal syndrome during heroin detoxification. No significant difference was found in withdrawal scores between the two groups at the beginning of the observation. Withdrawal scores showed a more marked drop in TEAS group than the control starting from the second day, and maintained at a lower level for the whole course of treatment. The area under the curve of withdrawal score in TEAS group was only 40% of that in the control (P < 0.001, two way repeated measures analysis of variance), and the requirement of buprenorphine was only 10% of that in the control. No adverse effects were observed in either group. CONCLUSION: TEAS of 2/100 Hz for 10 days in abrupt abstinence of the heroin addicts resulted in a marked reduction of the withdrawal syndrome as well as a reduced requirement for rescue opioids.

Genomewide analysis of rat periaqueductal gray-dorsal horn reveals time-, region- and frequency-specific mRNA expression changes in response to electroacupuncture stimulation.

Electroacupuncture (EA) has been widely applied for illness prevention, treatment or rehabilitation in the clinic, especially for pain management. However, the molecular events that induce these changes remain largely uncharacterized. The periaqueductal gray (PAG) and the spinal dorsal horn (DH) have been verified as two critical regions in the response to EA stimulation in EA analgesia. In this study, a genetic screen was conducted to delineate the gene expression profile in the PAG-DH regions of rats to explore the molecular events of the analgesic effect induced by low-frequency (2-Hz) and high-frequency (100-Hz) EAs. Microarray analysis at two different time points after EA stimulation revealed time-, region- and frequency-specific gene expression changes. These expression differences suggested that modulation of neural-immune interaction in the central nervous system played an important role during EA analgesia. Furthermore, low-frequency EA could regulate gene expression to a greater degree than high-frequency EA. Altogether, the present study offers, for the first time, a characterized transcriptional response pattern in the PAG-DH regions followed by EA stimulation and, thus, provides a solid experimental framework for future in-depth analysis of the mechanisms underlying EA-induced effects.

Dynamic changes of tyrosine hydroxylase and dopamine concentrations in the ventral tegmental area-nucleus accumbens projection during the expression of morphine-induced conditioned place preference in rats.

Our previous study demonstrated that morphine dose- and time-dependently elevated dopamine (DA) concentrations in the nucleus accumbens (NAc) during the expression of morphine-induced conditioned place preference (CPP) in rats. However, still unknown are how DA concentrations dynamically change during the morphine-induced CPP test and whether tyrosine hydroxylase (TH) activity in the ventral tegmental area (VTA) plays a vital role in this process. In the present study, we measured dynamic changes in TH and phosphorylated TH serine 40 (pTH Ser(40)) and pTH Ser(31) proteins in the VTA, and DA concentrations in the NAc at 5 min intervals during a 30 min morphine-induced CPP test. Rats that underwent morphine-induced CPP training significantly preferred the morphine-paired chamber during the CPP expression test, an effect that lasted at least 30 min in the drug-free state. DA concentrations in the NAc markedly increased at 15 min when the rats were returned to the CPP boxes to assess the expression of preference for the previously drug-paired chamber. DA concentrations then declined 2 h after the CPP test. TH and pTH Ser(40) levels, but not pTH Ser(31) levels, in the VTA were enhanced during the CPP test. These results indicated that TH and the phosphorylation of TH Ser(40) in the VTA may be responsible for DA synthesis and release in the NAc during the behavioral expression of conditioned reward elicited by a drug-associated context.

Electroacupuncture frequency-related transcriptional response in rat arcuate nucleus revealed region-distinctive changes in response to low- and high-frequency electroacupuncture.

Electroacupuncture (EA) has been clinically applied for treating different medical conditions, such as pain, strain, and immune diseases. Low- and high-frequency EAs have distinct therapeutic effects in clinical practice and experimental studies. However, the molecular mechanism of this difference remains obscure. The arcuate nucleus (Arc) is a critical region of the hypothalamus and is responsible for the effect of EA stimulation to remote acupoints. Gene expression profiling provides a powerful tool with which to explore the basis of physiopathological responses to external stimulus. In this study, using cDNA microarray, we investigated gene expressions in the rat Arc region induced by low-frequency (2-Hz) and high-frequency (100-Hz) EAs to two remote acupoints, zusanli (ST36) and sanyinjiao (SP6). We have found that more genes were differentially regulated by 2-Hz EA than 100-Hz EA (154 vs. 66 regulated genes/ESTs) in Arc, especially those related to neurogenesis, which was confirmed by qRT-PCR. These results demonstrate that the expression level of genes in the Arc region could be effectively regulated by low-frequency EA, compared with high-frequency EA, helping to uncover the mechanisms of the therapeutic effects of the low-frequency EA. Our results also indicate different-frequency EAs are spatially specific.

Multiple 100 Hz electroacupuncture treatments produced cumulative effect on the suppression of morphine withdrawal syndrome: Central preprodynorphin mRNA and p-CREB implicated.

Alleviating opiate withdrawal syndrome in addicts is a critical precondition to break away from drug and further to prevent reuse. Electroacupuncture (EA) was claimed to be effective for alleviating withdrawal syndrome, but the optimal protocol remained unclear. In the present study we found that (1) 100 Hz EA administered 12-24h after the last morphine injection suppressed the withdrawal syndrome in rats, multiple sessions of EA were more effective than single session, with the after-effect lasting for at least 7 days. (2) A down-regulation of preprodynorphin (PPD) mRNA level was observed in spinal cord, PAG and hypothalamus 60 h after the last morphine injection, which could be reversed by multiple sessions, but not a single session of EA. (3) Accompanied with the decrease of PPD mRNA level, there was an up-regulation of p-CREB in the three CNS regions, which was abolished by 100 Hz EA treatment. The findings suggest that down-regulation of p-CREB and acceleration of dynorphin synthesis in spinal cord, PAG and hypothalamus may be implicated in the cumulative effect of multiple 100Hz EA treatment for opioid detoxification.

Transcriptome profiling analysis reveals region-distinctive changes of gene expression in the CNS in response to different moderate restraint stress.

It is generally believed that temporary moderate stress to a living organism has protective and adaptive effects, but little is known about the responses of CNS to the moderate stresses at molecular level. This study aims to investigate the gene expression changes induced by moderate stress in CNS stress- and nociception-related regions of rats. Moderate restraint was applied to rats for 50 min and cDNA microarrays were used to detect the differential gene expression in different CNS regions. Transcriptome profiling analysis showed that at acute stage stress-related genes were up-regulated in arcuate nucleus; fight-or-flight behavior-related genes were up-regulated in periaqueductal gray, while nitric oxide and GABA signal transmission-related genes were up-regulated in spinal dorsal horn. In addition, immune-related genes were broadly regulated, especially at the late stage. These results suggested that specific genes of certain gene ontology categories were spatiotemporally regulated in specific CNS regions related to relevant functions under moderate external stimuli at acute stage, while immune response was broadly regulated at the late stage. The co-regulated genes among the three different CNS regions may play general roles in CNS when exposed to moderate stress. Furthermore, these results will help to elucidate the physiological processes involved in moderate stress in CNS.

Morphine-induced conditioned place preference in rats is inhibited by electroacupuncture at 2 Hz: role of enkephalin in the nucleus accumbens.

Our previous studies have demonstrated that morphine-induced conditioned place preference (CPP) can be inhibited by 2 Hz electroacupuncture (EA). This inhibition can be blocked by either the opioid receptor antagonist naloxone (i.p.) or lesion in the nucleus accumbens (NAc), providing evidence that endogenous opioid system in the NAc mediates the effects of EA. Here we report that 1) A single session of 2 Hz EA produced a significant increase of the content of enkephalin in the NAc of morphine-induced CPP rats, and this effect was stronger in three consecutive sessions of EA; 2) Intracerebroventricular injection of the mu-opioid receptor antagonist CTAP or delta-opioid receptor antagonist NTI, but not kappa-opioid receptor antagonist nor-BNI, dose-dependently reversed the inhibitory effects of 2 Hz EA on the expression of morphine-induced CPP; 3) Three consecutive sessions of 2 Hz EA up-regulated the mRNA level of preproenkephalin in the NAc of morphine-induced CPP rats. The results suggest that the inhibitory effects of 2 Hz EA on the expression of the morphine CPP is mediated by mu- and delta-, but not kappa-opioid receptor, possibly via accelerating both the release and synthesis of enkephalin in the NAc. These findings support the possibility of using 2 Hz EA for the treatment of opiate addiction.