Comparison of the analgesic properties of lipotropin C-Fragment and stabilized enkephalins in the rat

The C-Fragment of lipotropin (LPH 61–91) was shown to produce strong and long-lasting analgesia on intraventricular infusion in the rat. Its analgesic properties were compared with those of three synthetic derivatives of methionine enkephalin (LPH 61–65) which had been stabilized against enzymic degradation by blocking one or both termini with N-methyl and C-amide groups. C-Fragment was approximately 50 times more potent on a molar basis than N-methyl methionine enkephalin amide. The singly blocked pentapeptides, like methionine enkephalin, produced little more than transient analgesia. It was concluded that the analgesic properties of C-Fragment depend on the length and nature of the peptide chain rather than on the resistance of its N-terminal pentapeptide to degradation.     

Isolation of the C-fragment and C''-fragment of lipotropin from pig pituitary and C-fragment from brain.

Three novel peptides derived from lipotropin, the C-Fragment (residues 61-91), C'-Fragment (61-87) and N-Fragment (1-38), were isolated from pig pituitary, and the C-Fragment was shown to be present in brain. The experimental procedures developed for their isolation are described. The formation of each of the fragments involves enzymic cleavage of lipotropin at consecutive basic residues, with specificity identical with that involved in the activation of known prohormones. In brain assays C-Fragment exhibits a range of biological activities related to its ability to act as an inhibitory neurotransmitter.     

Lipotropin C-fragment has a COOH-terminal sequence with high intrinsic resistance to the action of exopeptidases.

The resistance of C-Fragment (lipotropin 61–91) to the action of carboxy-peptidase enzymes is shown to be due to the presence of consecutive lysine residues in the COOH-terminal sequence -Lys-Lys-Gly-Gln. A synthetic hexapeptide Ala-Tyr-Lys-Lys-Gly-Gln exhibited the same resistance, whereas the corresponding peptide with a single lysine was digested rapidly. The COOH-terminal stability of C-Fragment is thus a property of the primary structure and does not depend upon a specific molecular conformation. The significance of the stability is discussed in relation to the duration of biological activities exhibited by C-Fragment in the presence of brain exopeptidases.     

大鼠酒精躯体依赖及动机行为研究

目的：诱导大鼠产生酒精依赖,观察大鼠产生的躯体依赖及行为学改变。方法：20只雄性SD大鼠,其中饮酒组和对照组各10只。通过6％（v／v）酒精溶液作为饮酒组大鼠唯一饮水来源共28d,测量血酒精浓度。根据旷场行为、戒断症状和强迫游泳等方法来判断是否成功诱导大鼠产生酒精躯体依赖及动机行为改变。结果：整个实验过程饮酒组大鼠血酒精浓度没有发生明显变化。饮酒组大鼠旷场测试中水平活动量在饮酒第7d比对照组显著降低（P〈0．05）;垂直活动量在饮酒第7、14d比对照组显著降低（P〈0．05）;饮酒组大鼠戒断2-48h酒精戒断评分均显著高于对照组（P〈0．01）且评分在戒断第6h最高;戒断24、48h的大鼠在强迫游泳中绝对不动时间比对照组显著延长（P〈0．05）。结论：大鼠持续饮用6％（v／v）浓度酒精溶液可以诱导出大鼠对酒精的严重躯体依赖和抑郁状态,并抑制大鼠在新奇环境中的活动能力和动机行为。     

Oxytocin reduces intravenous heroin self-administration in heroin-tolerant rats

Intravenous self-administration of heroin was studied in experimentally naive rats, as compared to this behavior in animals rendered tolerant to heroin by multiple injections. The tolerant rats also exhibited mild signs of spontaneous and naloxone-precipitated heroin withdrawal. Self-administration behavior developed earlier in the tolerant rats. In heroin-naive rats, oxytocin treatment did not influence the acquisition of heroin self-administration behavior. In the tolerant rats, on the other hand, oxytocin decreased the acquisition of heroin self-administration. When maintenance of heroin self-administration was studied in the tolerant rats, graded doses of oxytocin (0.05, 0.5 and 5 micrograms s.c.) decreased heroin intake. This finding, which is in agreement with previous data indicating that oxytocin attenuates the development of tolerance to and dependence on narcotic analgesics, suggests that the neuropeptide reduced the reinforcing efficacy of heroin in the tolerant organism.     

大鼠吗啡依赖模型的建立

本实验采用剂量递增法,吗啡的给药剂量从10 mg/kg递增至80 mg/kg,用药5天停药,并经腹腔注射纳洛酮2 mg/kg催促后,实验观察大鼠主要行为学和植物神经症状表现.结果表明戒断症状明显,成功建立了大鼠吗啡依赖催促戒断模型.     

Is morphine dependence mediated exclusively by the Mu receptor?

Regional cerebral glucose utilization (RCGU) and behavior during naloxone precipitated withdrawal were studied in rats made dependent on either sufentanil, a highly selective mu opiate agonist, or morphine. Changes in RCGU during withdrawal in 23 of 24 anatomically related limbic and brainstem structures were indistinguishable between rats dependent on morphine or sufentanil. Linear regression analysis indicated that RCGU changes during withdrawal in these two groups were highly correlated (r=0.95). Withdrawal behaviors (autonomic signs of withdrawal, jumps, weight loss, and incidence of diarrhea) were likewise indistinguishable between the two groups. The correlations of both these measurements of dependence suggest that chronic occupation of mu opiate receptors alone is sufficient to produce dependence that is indistinguishable from morphine dependence.     

Behavioral Characterization of the Effects of Cannabis Smoke and Anandamide in Rats

Cannabis is the most widely used illicit drug in the world. Delta-9-tetrahydrocannabinol (Δ9-THC) is the main psychoactive component of cannabis and its effects have been well-studied. However, cannabis contains many other cannabinoids that affect brain function. Therefore, these studies investigated the effect of cannabis smoke exposure on locomotor activity, rearing, anxiety-like behavior, and the development of dependence in rats. It was also investigated if cannabis smoke exposure leads to tolerance to the locomotor-suppressant effects of the endogenous cannabinoid anandamide. Cannabis smoke was generated by burning 5.7% Δ9-THC cannabis cigarettes in a smoking machine. The effect of cannabis smoke on the behavior of rats in a small and large open field and an elevated plus maze was evaluated. Cannabis smoke exposure induced a brief increase in locomotor activity followed by a prolonged decrease in locomotor activity and rearing in the 30-min small open field test. The cannabinoid receptor type 1 (CB₁) receptor antagonist rimonabant increased locomotor activity and prevented the smoke-induced decrease in rearing. Smoke exposure also increased locomotor activity in the 5-min large open field test and the elevated plus maze test. The smoke exposed rats spent more time in the center zone of the large open field, which is indicative of a decrease in anxiety-like behavior. A high dose of anandamide decreased locomotor activity and rearing in the small open field and this was not prevented by rimonabant or pre-exposure to cannabis smoke. Serum Δ9-THC levels were 225 ng/ml after smoke exposure, which is similar to levels in humans after smoking cannabis. Exposure to cannabis smoke led to dependence as indicated by more rimonabant-precipitated somatic withdrawal signs in the cannabis smoke exposed rats than in the air-control rats. In conclusion, chronic cannabis smoke exposure in rats leads to clinically relevant Δ9-THC levels, dependence, and has a biphasic effect on locomotor activity.     

Effects of stress and alcohol consumption on emotional/motivational behavior of rats and their first generation offspring

Neurophysiological manifestations of formation of the dependence on ethanol under stress conditions were studied in rats and their first generation offspring. In chronic experiments on 32 male rats, emotional stress was modeled (induction by nociceptive electrocutaneous stimulation), and an increased addiction to ethanol was formed. In these animals and in the first generation of their offspring (obtained from the above males and intact females), we studied emotional/motivational behavior, recorded the mass electrical activity from different brain structures, and measured the arterial pressure. In stressed rats, which acquired attraction to ethanol, negative emotional responses became transformed into positive, and behavioral and electrographic manifestations of the seizure activity developed. In the first generation offspring, the pattern of central rearrangements in the mechanisms of emotional/motivational behavior was to a great extent similar to that in parent rats.     

Plasticity of GABA<Subscript>A</Subscript> Receptors in Brains of Rats Treated with Chronic Intermittent Ethanol

The study of alcohol dependence mechanisms has been aided by work in rodents, where regimens of intermittent chronic administration with repeated episodes of intoxication and withdrawal can be coupled with controlled timing of in vitro studies and the possibility of relating them to behavior. The chronic intermittent ethanol (CIE) model in the rat has been found to be a good model of human alcohol dependence, showing persistent signs of withdrawal and self-administration. Studies in CIE rats suggest that plastic changes in GABA-mediated inhibition involving the GABA_A receptor system may be responsible for the behavioral alterations. Here we summarize a combination of evidence that the alcoholic rat CIE model demonstrates changes in GABA_A receptor subunit levels, in receptor localization, and in physiology and pharmacology, leading to alterations in behavior that contribute to the hyperexcitable alcohol withdrawal state (anxiety, insomnia, seizure susceptibility) and alcohol dependence. Special Issue dedicated to Dr. Simo S. Oja     

Maternal separation and early stress cause long‐lasting effects on dopaminergic and endocannabinergic systems and alters dendritic morphology in the nucleus accumbens and frontal cortex in rats

A considerable amount experimental studies have shown that maternal separation (MS) is associated with adult offspring abnormal behavior and cognition disorder. Accordingly, this experimental procedure has been proposed as a predictor for alcohol and drug dependence based on the neurodevelopmental soon after birth. Endocannabinoid system (eCBs) has been implicated in reward processes, including drug abuse and dependence. MS and associated stress causes changes in the eCBs that seem to facilitate alcohol consumption. In this study, we seek to evaluate potential morphological changes in neurons of the frontal cortex (FCx) and nucleus accumbens (NAcc), in the expression of receptors and enzymes of the endocannabinoid and dopamine systems and in second messengers, such as Akt, in adult rats subjected to MS and early stress (MS + ES; 2 × 180 min daily) vs. nonseparated rats (NMS). Results showed that MS + ES induces higher D2R expression and lower D3R, FAAH, and MAGL expression compared with NMS rats. Alterations in total dendritic length were also detected and were characterized by increases in the NAcc while there were decreases in the FCx. We believe MS + ES‐induced changes in the dopaminergic and endocannabinergic systems and in the neuronal microstructure might be contributing to alcohol seeking behavior and, potential vulnerability to other drugs in rats. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 819–831, 2016     

Independence of changes in contraction properties and myofibrillar ATPase activity of denervated mammalian muscle on length of nerve stump.

1. Contractile properties of the fast extensor digitorum longus of one-month-old rats and of the fast peroneus longus muscles of adult rabbits were studied in vitro at 36 degrees C after nerve section close to the muscle. Changes in contraction properties (prolongation) are not observed until 48 hours after denervation in the rat and 14-30 days in the rabbit. 2. At no period after denervation are differences in twitch isometric contraction properties dependent on the length of the sectioned nerve stump. This lack of dependence of contractile behavior after denervation is in contrast to many metabolic changes which show a clear dependence on the length of the nerve stump. 3. It is concluded that the onset of denervation changes in contractile behavior are related to the loss of nerve-impulse activity, while the transient early metabolic changes are related to changes of fast axoplasmic flow, initiated after nerve section and therefore dependent on length of sectioned nerve stump.     

Nandrolone modulates the non-opioid and opioid analgesia and tolerance/dependence: role of sexual dimorphism

The aim of this investigation was to study the effect of the doping steroid nandrolone on metamizol and morphine-induced analgesia and tolerance/dependence in rats. Nandrolone per se did not change the basal nociceptive thresholds in both sexes. It diminished the analgesic effect of metamizol in females, revealed by tail flick test, and males, revealed by paw pressure and hot plate tests. In general, the action of nandrolone was to decrease the morphine-induced analgesia in female and male rats. This was strongly manifested by paw pressure and tail flick tests in male, and tail flick tests in female animals. Nandrolone slowed the development of opioid tolerance/dependence. It aggravated the withdrawal syndrome in the females and invigorated aggression in the males. The data provide evidence that anabolic steroid nandrolone might decrease the analgesic action of metamizol or morphine. The doping steroid could modulate opioid tolerance/dependence and the aggressive behavior in a gender dependent manner. The action of nandrolone is most likely due to profound long-term effects on the central nervous system and might be a gateway to addiction of other drugs of abuse.     

Behavioral dependence on phencyclidine in rats

The abuse of PCP continues to be an important medical problem in many urban areas. The probability that dependence on PCP may contribute to its compulsive use and relapse is supported by animal studies demonstrating its dependence liability. In the present study, five rats were housed in operant chambers and trained to respond on a lever under a fixed-ratio 30 schedule of food presentation. They obtained all their daily food during four 30-min response periods occurring every 6 hr. After stable baselines of behavior were established the rats were injected with PCP (3.0-7.5 mg/kg/injection), i.p., 1 hr before each response session for 7-10 days. Following chronic dosing, the drug injections were replaced with saline injections for 10 days. Disruptions in behavior were observed upon cessation of relatively brief chronic exposure to PCP (as little as 7 days) and at relatively low doses (5.6 mg/kg/6 hr = 22.4 mg/kg/day). The behavioral disruption was not accompanied by overt signs of abstinence and persisted for up to 48 hr.     

JWA regulates chronic morphine dependence via the delta opioid receptor

Opioid dependence is correlated with the adaptive changes at the cellular level following chronic opioid use, and believed to be the main cause for the relapse of drug taking behavior of addicts. Despite decades of intensive studies, the underlying mechanisms of morphine dependence are still unclear. Here, we present evidence that JWA was induced by chronic morphine treatment in specific brain regions, and knockdown of JWA expression significantly reduced the withdrawal response to chronic morphine treatment in rats. We further demonstrated that the morphine induced DOR expression, while activation of DARPP-32 and MAP kinase was suppressed by JWA knockdown. Through an in vitro cell model of chronic morphine exposure, we also found that JWA is required for maintaining the stability of DOR via the ubiquitin–proteasome pathway. These observations suggest that JWA is directly involved in the regulation of chronic morphine dependence.     

Differential maintenance of penile responses and copulatory behavior by gonadal hormones in castrated male rats

Sexually experienced male rats were castrated and immediately received implants of Silastic tubing containing either testosterone (T), dihydrotestosterone (DHT), estradiol (E), or nothing (blank). The ability of these hormone treatments to maintain precastration levels of copulatory behavior and ex copula penile responses was assessed for 40 days after castration. Throughout the study T- and E-treated males, but not males with DHT or blank implants, maintained normal copulatory behavior. In contrast males treated with T and DHT, but not E or blanks, maintained penile responses ex copula. In blank-treated males, penile-response latencies increased more rapidly than did intromission latencies. These results, together with those of previous studies, appear to rule out a role for estradiol and reinforce the role of androgens in the activation of rats' penile-response potential ex copula. Similarly, the results support the conclusion that in castrated male rats estradiol treatment is sufficient for the activation of masculine copulatory behavior, and that the penile actions necessary for intromission are not dependent on androgen. Thus, the evocability of penile actions and their relative androgen dependence are context sensitive.     

Cocaine withdrawal produces behavioral disruptions in rats

There is currently no laboratory or clinical evidence from animal or human studies documenting a withdrawal syndrome associated with cocaine dependence, although many users report that withdrawal disturbances are responsible for their repeated use of the drug. In the present study rats self-administered i.v. cocaine and a sweetened drinking solution. When cocaine access was terminated there was a marked suppression in operant behavior reinforced by the sweetened solution, and this withdrawal disruption was immediately reversed when cocaine was reinstated. There were no physical signs of withdrawal, and food intake increased when cocaine was withdrawn. The results suggest that sensitive behavioral tests reveal aspects of drug dependence that may account for persistent abuse.     

The AQP2 water channel: Effect of vasopressin treatment,microtubule disruption,and distribution in neonatal rats

Aquaporin 2 is a collecting duct water channel that is located in apical vesicles and in the apical plasma membrane of collecting duct principal cells. It shares 42% identity with the proximal tubule/thin descending limb water channel, CHIP28. The present study was aimed at addressing three questions concerning the location and behavior of the AQP2 protein under different conditions. First, does the AQP2 channel relocate to the apical membrane after vasopressin treatment? Our results show that AQP2 is diffusely distributed in cytoplasmic vesicles in collecting duct principal cells of homozygous Brattleboro rats that lack vasopressin. In rats injected with exogenous vasopressin, however, AQP2 became concentrated in the apical plasma membrane of principal cells, as determined by immunofluorescence and immunogold electron microscopy. This behavior is consistent with the idea that AQP2 is the vasopressin-sensitive water channel. Second, is the cellular location of AQP2 modified by microtubule disruption? In normal rats, AQP2 has a mainly apical and subapical location in principal cells, but in colchicine-treated rats, it is distributed on vesicles that are scattered throughout the entire cytoplasm. This is consistent with the dependence on microtubules of apical protein targeting in many cell types, and explains the inhibitory effect of microtubule disruption on the hydroosmotic response to vasopressin in sensitive epithelia, including the collecting duct. Third, is AQP2 present in neonatal rat kidneys? We show that AQP2 is abundant in principal cells from neonatal rats at all days after birth. The detection of AQP2 in early neonatal kidneys indicates that a lack of this protein is not responsible for the relatively weak urinary concentrating response to vasopressin seen in neonatal rats.     

Aggressive behavior and radiosensitivity in rats'

This paper aims to present the study of rats' individual radiosensitivity dependence on their individual aggressiveness. On total irradiation in sublethal doses (1.0, 1.5 and 3.5 Gy) and in doses close to LD50/30 (6, 7 and 8 Gy) there was investigated comparative radiosensitivity of non-aggressive and aggressive rats of Wistar line, as well as that of non-aggressive individuals during provoked aggressiveness by means of blocking serotonin synthesis with intraperitoneal (i/p) injection of 400 mg/kg of parachlorphenylalanine (pCPA). Muricidity served as a criterion for aggressiveness and as a criterion of radiosensitivity--cumulative function of survival, the changes of behavior in "Open feald", serotonine and catecholamine content in various brain structures and the dose dependence on the radiation modification of muricidity. It has been found that after 1 Gy total X-irradiation the rats do not lose aggressiveness. Nevertheless the ethalogical parameters change in considerable degree. In the doses of 1.5 and 3.5 Gy muricidity is eluminated in 15-18 and 5-9 days, correspondingly. I/p injection of pCPA after the elimination of aggressiveness provokes transient muricidity in the same terms and duration as it is in case of non-aggressive rats. The elimination of muricidity is associated with changes in content and distribution of biogenic amines in various structures of brain, as well as with reduction of locomotor and reference-research activity, on the one hand and with an increase of emotionality and stereotype activity, on the other hand. After X-irradiation in 6, 7 and 8 Gy the regression coefficients of the dependence of functions type of survival on irradiation dose in aggressive rats is significantly reliable both in comparison with non-aggressive rats and animals with provoked aggressiveness. The change of mortality-rate per unit of changing irradiation dose not depend on blocking of serotonin synthesis, which deficit is one of the distinct determinant of aggressiveness, on the one hand, and higher radiosensitivity, on the other hand. The obtained data allow to suppose that elimination of muricidity after the irradiation of rats in the sublethal doses is conditioned not only by the consequenses of radiation damage of neurobiological structures responsible for the organization of aggressive behavior but the activation of serotonergic system in the process of restitution after radiation trauma. On the other hand, higher radiosensitivity of aggressive rats compared with non-aggressive ones is connected with low serotonin content, thiols and some other biologically active substances which are endogenous radioprotectors determining individual radioresistance.     

多巴胺D2受体拮抗剂、激动剂对SD大鼠条件化位置偏好的影响

通过慢性吗啡处理方式建立起SD大鼠吗啡依赖的条件化位置偏好(CPP)模型,用行为学手段研究多巴胺(DA)D2受体拮抗剂及激动剂对SD大鼠CPP的影响,探讨眶额叶DAD2受体在阿片精神依赖中的作用。通过腹腔注射吗啡同环境因素相结合,建立大鼠吗啡依赖的CPP模型;采用局部脑内微量注射法向额叶注射DAD2受体拮抗剂或激动剂或盐水(对照组),以得到SD大鼠在戒断期间的CPP的时间数据。CPP显示DAD2受体拮抗剂组与对照组相比,从戒断第2天起,前者表现出更明显的CPP增加现象,差异显著(P<0·05)。而DAD2受体激动剂组与对照组相比无显著差异(P>0·05)。采用腹腔小剂量注射吗啡,成功地建立了吗啡依赖SD大鼠的CPP模型;眶额叶微量注射DAD2受体拮抗剂增加了CPP时间,提示眶额叶多巴胺系统在吗啡依赖的过程中有着较为重要的作用;也提示了对于已经成瘾的动物,损伤其眶额叶,会使药物渴求增强。因而提示对于药物依赖患者进行手术干预治疗要极其慎重。