Sexual dimorphism in the brain underlies behavioral differences between sexes. The bed nucleus of the stria terminalis (BNST) is a complex nucleus that differs between males and females, but the sexual dimorphism in cytoarchitecture and the connectome of its oval subdivision (ovBNST) remains largely unexplored. By combining snRNA-seq and transgenic labeling, we found a higher density of ovBNST proenkephalin (ovBNST^PENK) neurons in male than female mice. Anatomically, we virally mapped the efferents and afferents of ovBNST^PENK neurons, finding reciprocally dimorphic connections with the hypothalamus and striatum. Gene enrichment analysis suggests that ovBNST^PENK neurons are modulated by the upstream dopamine pathway. Functionally, by applying caspase-3-mediated depletion of ovBNST^PENK neurons, we found that loss of these neurons enhanced locomotor activity in male but not female mice, without altering the anxiety-like phenotypes in either sex. Our study may pave the way for a better understanding of the anatomical and functional profiles of ovBNST^PENK neurons from a sexually dimorphic perspective.
Animals ; Male ; Female ; Septal Nuclei/physiology* ; Sex Characteristics ; Neurons/physiology* ; Enkephalins/metabolism* ; Mice ; Mice, Transgenic ; Protein Precursors/metabolism* ; Mice, Inbred C57BL ; Neural Pathways/physiology*

OBJECTIVETo observe analgesic effect of electroacupuncture ( EA) on rats with chronic inflammatory pain and its regulatory mechanism on ispilateral dorsal root ganglion (DRG) and spinal dorsal horn (SDH) Mas-related G protein-coupled C receptor (MrgprC).

METHODSTotally 40 healthy male SD rats were divided into 4 groups according to random number table, i.e., the normal (N) group, the model (M) group, the acupuncture (Acu) group, the EA group, 10 rats in each group. The model of chronic inflammatory pain was established by subcutaneous injecting 0. 1 mL complete Freund's adjuvant (CFA) into right hind paw. Paw withdrawal thresholds (PWTs) were measured before modeling, at day 1, 3, 5, 7, and after CFA injection, respectively. Expression levels of MrgprC in ispilateral DRG and SDH were detected by Western blot. The content of bovine adrenal medulla 22 (BAM22) in SDH was detected by immunohistochemical assay.

RESULTSCompared with N group at each time point, PWTs significantly decreased in M group (P <0. 01). Compared with M group, PWTs significantly increased at day 5 of EA and after EA in EA group (P < 0.05, P < 0.01). Compared with Acu group at each time point, post-EA PWTs significantly increased in the EA group (P < 0.05). Compared with N group, expression of MrgprC in ispilateral DRG and ratio of BAM22 positive cells in ispilateral SDH increased in M group (P < 0.01). Compared with M group, expression of MrgprC in ispilateral DRG and ratio of BAM22 positive cells in ispilateral SDH increased in the EA group (P < 0.05).

CONCLUSIONEA had favorable analgesic effect on chronic inflammatory pain induced by CFA, and its mechanism might be possibly associated with up-regulating MrgprC expression in ispilateral DRG and BAM22 content in ispilateral SDH.

Analgesia ; Animals ; Electroacupuncture ; Enkephalins ; metabolism ; Freund's Adjuvant ; Ganglia, Spinal ; drug effects ; Inflammation ; chemically induced ; drug therapy ; Male ; Pain Management ; methods ; Peptide Fragments ; metabolism ; Posterior Horn Cells ; drug effects ; Random Allocation ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo explore the analgesic mechanism of small knife needle for treating transverse process syndrome of the third vertebra (TPSTV) by observing peripheral and central changesof β-endorphin (β-EP) and enkephalin (ENK) contents.

METHODSTotally 30 Japanese white big-ear rabbits of clean grade were divided into 5 groups according to random digit table, i.e., the normal control group, the model group, the small knife needle group, the electroacupunture (EA) group, and the small knife needle plus EA group, 6 in each group. The TPSTV model was established by inserting a piece of gelatin sponge into the left transverse process of 3rd lumbar vertebrae. Rabbits in the small knife needlegroup were intervened by small knife needle. Those in the EA group were intervened by EA at bilateralWeizhong (BL40). Those in the small knife needle plus EA group were intervened by small knife needleand EA at bilateral Weizhong (BL40). Contents of β-EP and ENK in plasma, muscle, spinal cord, and hypothalamus were determined after sample collection at day 28 after modeling.

RESULTSCompared with the normal control group, contents of β-EP and ENK in plasma and muscle increased significantly, and contents of β-EP and ENK in spinal cord and hypothalamus decreased significantly in the model group (P < 0.05, P < 0.01). Contents of β-EP and ENK approximated normal levels in the three treatment groups after respective treatment. Compared with the model group, the content of β-EP in muscle decreased, and contents of β-EP and ENK in hypothalamus increased in the three treatment groups after respective treatment (P < 0.05). There were no significant difference among the three treatment groups (P > 0.05).

CONCLUSIONSSmall knife needle treatment and EA had benign regulation on peripheral and central β-EP and ENK in TPSTV rabbits. Small knife needle treatment showed better effect than that of EA.

Acupuncture Points ; Animals ; Electroacupuncture ; Enkephalins ; metabolism ; Hypothalamus ; metabolism ; Lumbar Vertebrae ; pathology ; Muscle, Skeletal ; metabolism ; Needles ; Rabbits ; Random Allocation ; Spinal Cord ; metabolism ; Spinal Diseases ; therapy ; beta-Endorphin ; metabolism

OBJECTIVETo observe the intervention of electroacupuncture (EA) with different current frequencies and treatment frequencies on pain thresholt in rats with bone-cancer pain, so as to optimize treatment parameters of EA against bone cancer pain; and by measuring gene expression of opioid receptor and precursor in different tissues to preliminarily explore the possible mechanism of EA against bone cancer pain.

METHODSNinety healthy female SD rats were randomly divided into a control group, a model group, EA groups (6 subgroups according to different frequencies) and a sham EA group, ten rats in each one. Rats in the control group were injected with 10 µL of amicrobic phosphate buffer solution (PBS) into tibial cavity; rats in the remaining groups were injected with Walker 256 cancer cells to establish model of bone-cancer pain. No treatment was given to rats in the control group and model group; rats in the EA groups were treated with EA at bilateral "Housanli" (ST 36) and "Genduan" with 3 different current frequencies (2 Hz, 100 Hz and 2 Hz/100 Hz), once a day and once every other day, 30 min per treatment (1mA for 15 min, 2 mA for 15 min); rats in the sham EA group were treated with identical acupoints as the EA group, but the acupoints were needled subcutaneously and EA was connected with power off. All the treatment was given for 14 days. Dynamic plantar aesthesiometer was applied to measure the paw withdrawal thresholds (PWTs) of the affected side before the model establishment, 6d, 8d, 10d, 12d, 14d, 16d, 18d, and 20d after model establishment. The mRNA expressions of µ-opioid receptor (MOR), κ-opioid receptor (KOR), δ-opioid receptor (DOR), proopiomelanocortin (POMC) and prodynorphin (PDYN) in dorsal root ganglion (DRG) and lumbar spinal cord dorsal horn (SCDH) of L4-L6 of the affected side were detected by PCR method.

RESULTSThere were no differences in PWTs among all groups before model establishment (P>0. 05). Each time point after model establishment, PWTs in model group were obviously lower than those in the control group (all P<0. 01). Compared with the model group, PWTs in each EA subgroup were all increased (all P<0.05), but the differences at different time points were not significant among EA subgroups (P>0.05). The mRNA expressions of MOR, KOR, POMC, and PDYN in L4-L6 DRG in the 2 Hz/100 Hz II group were significantly higher than those in model group (P<0. 05, P<0. 01), while the mRNA expressions of MOR, KOR, DOR, POMC and PDYN in SCDH were not different compared with the model group (P>0. 05).

CONCLUSIONEA treatment has obvious analgesic effect on bone-cancer pain, however, its effect is not related with current frequency and treating frequency. EA against bone-cancer pain may be related with increasing the mRNA expression of some peripheral opioid receptors and precursor.

Acupuncture Analgesia ; instrumentation ; methods ; Acupuncture Points ; Animals ; Bone Neoplasms ; complications ; Electroacupuncture ; instrumentation ; methods ; Enkephalins ; metabolism ; Female ; Ganglia, Spinal ; metabolism ; Humans ; Pain ; etiology ; genetics ; metabolism ; Pain Management ; instrumentation ; methods ; Protein Precursors ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid ; genetics ; metabolism

OBJECTIVETo explore the central mechanism of moxibustion on analgesic effect.

METHODSMale Wistar rats were screened by pain threshold value before making model, and 48 rats whose pain threshold was (250 +/- 25) g were selected. Twelve male Wistar rats were randomly selected as a normal group. For the rest rats the rheumatoid arthritis (RA) model was duplicated by raising in a windy, cold and wet environment combined with injection of Freund's complete adjuvant (FCA), and then they were randomly divided into a model group, a moxibustion group and a moxa volatile oil group, 12 rats in each group. The moxibustion and the moxa volatile oil igroup were treated with moxibustion and moxa volatile oil at "Shenshu"(BL 23) and "Zusanli"(ST 36), respectively, for 15 days. No interventions were added on the model group and the normal group. The pain threshold in Iinjured foot and the expression of hypothalamic POMC mRNA and PDYN mRNA in rats were observed.

RESULTSCompared with the normal group, the pain threshold and the expression of hypothalamic POMC mRNA and PDYN mRNA in the model group were increased (all P < 0.01). Compared with the model group, the pain threshold and the expression of hypothalamic POMC mRNA and PDYN mRNA in the moxibustion group were increased significantly (all P < 0.01), but no statistically significance in the moxa volatile oil group (P > 0.05). Compared with the moxa volatile oil group, the above-mentioned observative indices in moxibustion group were all increased significantly (all P < 0.01).

CONCLUSIONMoxibustion has obvious analgesic effect and its mechanism may be related to the increasing expression of hypothalamic POMC and PDYN mRNA through the warming effect of moxibustion.

Animals ; Arthritis, Rheumatoid ; genetics ; metabolism ; therapy ; Enkephalins ; genetics ; metabolism ; Humans ; Hypothalamus ; metabolism ; Male ; Moxibustion ; Pro-Opiomelanocortin ; genetics ; metabolism ; Protein Precursors ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Wistar

OBJECTIVETo provide a sound cell source for further ex-vivo gene therapy for chronic pain, we attempt to develop an immortalized rat astrocyte cell line that expresses enkephalin regulated by doxycycline.

METHODSRetrovirus infection method was employed to develop an immortalized rat astrocyte cell line that could express enkephalin regulated by doxycycline. The hPPE gene expression level of immoralized astroyte cells (IAC)/ hPPE was detected by RT-PCR, indirect immunofluorescence staining and radioimmunoassay.

RESULTSIAC carrying Tet-on system transfected with preproenkephalin gene could secrete enkephalin that was regulated by doxycycline in a dose-dependent manner and hPPE gene activation could be repeated in on-off-on cycles through administration or removal of doxycycline.

CONCLUSIONAn immortalized rat astrocyte cell line that secrete enkephalin under the control of doxycycline is established successfully, which provides a research basis for transgenic cell transplantation for analgesia.

Animals ; Anti-Bacterial Agents ; pharmacology ; Astrocytes ; Cell Line, Transformed ; Chronic Disease ; Doxycycline ; pharmacology ; Enkephalins ; genetics ; metabolism ; Genetic Therapy ; methods ; Genetic Vectors ; Neurotransmitter Agents ; genetics ; metabolism ; Pain Management ; Protein Precursors ; genetics ; metabolism ; Rats ; Retroviridae ; genetics

Bovine adrenal medulla 22 (BAM22), an endogenous opioid peptide, is one of the cleavage products of proenkephalin A. It potently activates opioid receptors and sensory neuron-specific receptor (SNSR). The present study was aimed at investigating the effect of BAM22 on morphine tolerance. Intrathecal (i.t.) administration of morphine for 7 d produced morphine tolerance in rats. Then the rats were divided into three groups in which morphine, saline or BAM22 were administered i.t., respectively, on day 8, and morphine was given to all of the animals on day 9. It was found that morphine administered on day 9 resumed antinociceptive effects in BAM22 group, but not in saline or morphine group. The potency of morphine in BAM22 group was 48.5% of the maximal possible effect (MPE) detected by paw withdrawal test and the antinociception persisted for approximately 1 h. Following the similar treatment, morphine administered on day 9 reduced nocifensive behaviors by 3.2 min and 24 min in BAM22 group in the first and second phases, in the formalin test, respectively. The decreases were 45% and 82% of the corresponding values observed in saline group. Furthermore, following the treatment with BAM22 (10 nmol) on day 8 in morphine-tolerance rats, morphine administered on day 9 decreased the expressions of the heat-evoked c-Fos-like immunoreactivity (FLI) protein by approximately 80% in laminae I-II, III-IV and V-VI in the spinal cord at L4-L5 compared with that in saline or morphine group. The present study provided evidence at behavioral and cellular levels showing that BAM22 resumed antinociception of morphine. The results that the reversal effect of BAM22 on morphine tolerance was more efficient in persistent pain model than in acute pain may indicate that BAM22 differentially modulates morphine tolerance. The present study suggests that SNSR is involved in the modulation of morphine tolerance.
Animals ; Drug Tolerance ; Enkephalins ; pharmacology ; Morphine ; pharmacology ; Pain ; drug therapy ; Peptide Fragments ; pharmacology ; Rats ; Receptors, G-Protein-Coupled ; metabolism

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.
Animals ; Enkephalins ; genetics ; metabolism ; Formaldehyde ; Gene Transfer Techniques ; Genetic Therapy ; Genetic Vectors ; administration & dosage ; Herpesvirus 1, Human ; genetics ; Humans ; Injections, Spinal ; Male ; Nociceptors ; physiology ; Pain ; chemically induced ; Pain Management ; Protein Precursors ; genetics ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley

AIMTo investigate the changes of enkephalin mRNA and prodynorphin mRNA gene expression in rat brain regions with chronic immobilization stress and the influence of Chinese herbs.

METHODSWe copied the rat model of chronic immobilization stress (3 h daily , repeated 7 d or 21 d), and primers of enkephalin or prodynorphin were respectively added for RT-PCR reaction (BETA-actin as inner contrast). Gel image analysis system was used to scan and analyze and odds of optical density of target gene and inner contrast strip were taken as quasi-quantified data.

RESULTSProdynorphin mRNA expression in hippocampus markedly increased in 7 d group (P < 0.01), while enkephalin mRNA prodynorphin mRNA expression in hippocampus markedly increased in 21 d group (P < 0.01). All three recipes were able to decrease the gene expression of prodynorphin mRNA (P < 0.01), and xiaoyao powder as well as sijunzi soup were able to decrease the gene expression of enkephalin mRNA (P < 0.01).

CONCLUSIONThe effects of xiaoyao powder on the above two gene expression was better than jinkuishenqi pill.

Animals ; Drugs, Chinese Herbal ; pharmacology ; Dynorphins ; metabolism ; Enkephalins ; metabolism ; Gene Expression ; Hippocampus ; drug effects ; metabolism ; Male ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley ; Restraint, Physical ; Stress, Physiological

BACKGROUND: Parkinson's disease is the primary degenerative disease characterized by rigidity, bradykinesia and resting tremor. Even though dopaminergic cell loss of the substantia nigra compacta is a main pathogenesis of the Parkinson's disease, the striatal dysfuntion is major pathophysiology of pakinsonian symptoms. METHODS: Since gene expression profile can explain the symptomatic varieties of Parkinson's disease, gene expression was investigated in the striatum of rat brain after lesioning of unilateral substantia nigra compacta with 6-hydroxydopamine using cDNA microarray technique for the first time. The expression patterns of 5,200 rat brain cDNAs were screened and clustered according to the function of gene. The expression patterns of enkephaline and substance-P mRNA were also studied for validation of animal preparation. RESULTS: Various genes involved in apoptosis, cytokines. cytoskeletal molecules, neurotrophic factors, receptors, intracellular Ca2+ metabolism, signal transduction, stress protein, cell cycle regulator protein, and expressed sequence tags(EST) have shown significant expression changes. CONCLUSIONS: These gene expressions provide the global assessment of the processes involved in secondary change of striatum afforded by dopaminergic denervation at molecular levels.
Animals ; Apoptosis ; Brain ; Cell Cycle ; Cytokines ; Denervation ; DNA, Complementary ; Enkephalins ; Gene Expression ; Hypokinesia ; Metabolism ; Models, Animal* ; Nerve Growth Factors ; Oligonucleotide Array Sequence Analysis ; Oxidopamine* ; Parkinson Disease ; Rats* ; RNA, Messenger ; Signal Transduction ; Substantia Nigra ; Transcriptome ; Tremor