Increased anxiety-related behavior in mice deficient for metabotropic glutamate 8 (mGlu8) receptor

Pre-synaptic metabotropic glutamate (mGlu) receptors modulate neuronal excitability by controlling glutamate and gamma-aminobutyric acid (GABA) release. The mGlu8 receptor is predominantly found in pre-synaptic terminals and its expression is highly restricted. To study the role of this receptor, mGlu8 receptor-deficient mice were generated. Here we report that na?ve mGlu8 receptor-deficient mice showed increased anxiety-related behavior in the elevated plus maze in low illumination conditions (red light). Open arm avoidance and risk assessment behavior were both significantly increased in mutant mice. Increased stressfulness of the testing conditions abolished this behavioral difference. Fluorescent light or prior restraint stress decreased the open arm activity of wild-type mice, while the open arm activity of mutant mice was essentially unaffected, leading to similar values in both strains. The total number of arm entries or closed arm entries was not significantly different between strains, indicating that the lack of mGlu8 receptor does not affect locomotor activity. No gross behavioral changes, or changes in the function of the autonomic nervous system or somatomotor systems were observed in mutant mice. Moreover, no significant differences in seizure susceptibility were detected between strains. Our results suggest that mGlu8 receptor may play a role in responses to novel stressful environment.     

Role of D1-like receptors in amphetamine-induced behavioral sensitization: a study using D1A receptor knockout mice

RATIONALE: The role played by D(1)-like receptors in amphetamine-induced behavioral sensitization has been examined using both the D(1)-like receptor antagonist, SCH 23390, and the D(1A) receptor knockout mouse (i.e. D(1A)-deficient mice). Studies using these two approaches have provided conflicting evidence about the importance of D(1)-like receptors for amphetamine-induced behavioral sensitization. OBJECTIVE: The purpose of the present study was to determine: (a) whether D(1A)-deficient mice exhibit amphetamine-induced locomotor sensitization after 3 and 17 drug abstinence days, and (b) whether SCH 23390, which binds to both D(1A) and D(1B) receptor subtypes, blocks development of amphetamine sensitization in wild-type and D(1A)-deficient mice. METHODS: In the first experiment, adult wild-type and D(1A)-deficient mice were injected with amphetamine (0, 1, 2, 4, or 8 mg/kg, IP) for 7 consecutive days. In the second experiment, wild-type and D(1A)-deficient mice were pretreated with SCH 23390 (0, 0.15, or 0.5 mg/kg, IP) 30 min prior to being injected with amphetamine (0 or 8 mg/kg, IP). After each daily amphetamine injection, mice were placed in activity chambers where distance traveled (i.e. horizontal locomotor activity) was measured for 60 min. On the test days, which occurred after 3 or 17 drug abstinence days, mice were injected with 1 mg/kg amphetamine and locomotion was measured for 120 min. RESULTS: Both wild-type and D(1A)-deficient mice exhibited amphetamine-induced locomotor sensitization. Pretreatment with 0.5 mg/kg SCH 23390 blocked the development of locomotor sensitization in wild-type mice, but did not alter the sensitized responding of D(1A)-deficient mice. CONCLUSIONS: It appears that D(1)-like receptors are necessary for the development of amphetamine sensitization in wild-type mice, while neither the D(1A) nor D(1B) receptor subtypes are necessary for the amphetamine-induced locomotor sensitization of D(1A)-deficient mice. A possible explanation for these conflicting results is that D(1A)-deficient mice may have a compensatory mechanism (not involving D(1B) receptors) that allows them to exhibit amphetamine-induced behavioral sensitization in the absence of the D(1A) receptor.     

Alteration of voluntary ethanol and saccharin consumption by the neurosteroid allopregnanolone in mice

RATIONALE: The neurosteroid 3 alpha-hydroxy-5 alpha-pregnan-20-one (allopregnanolone, ALLOP) is a positive modulator of gamma-aminobutyric acid type A (GABA(A)) receptors. Recent findings indicate that ethanol (EtOH) and ALLOP share common mechanisms of action and that ALLOP may modulate some of EtOH's abuse-related effects. OBJECTIVES: The present studies investigated whether ALLOP pretreatment altered voluntary EtOH consumption in male and female C57BL/6J mice, and voluntary saccharin and quinine consumption in male C57BL/6J mice. METHODS: Mice had access to two drinking tubes containing water versus 5% or 10% (v/v) EtOH or a tastant for 2 h each day at the beginning of the dark cycle. Following establishment of stable consumption, animals received 2 days of vehicle followed by 3 days of ALLOP injections (0, 3.2, 10, or 17 mg/kg, IP), immediately prior to EtOH or tastant access. RESULTS: Prior to injection, the 2-h baseline dose of the 10% EtOH solution consumed was 1.31 g/kg (expt 1) or 2.46 g/kg (expt 3) for male and 2.21 g/kg (expt 2) for female mice. Baseline intake of the 5% EtOH solution was 0.60 g/kg for males and 0.75 g/kg for females (expt 5). In males, ALLOP administration significantly and dose-dependently increased consumption of both EtOH solutions during the first hour of availability without affecting water intake. In females, ALLOP did not significantly alter EtOH consumption. Lastly, ALLOP significantly increased saccharin, but not quinine, consumption in males (females were not tested). CONCLUSIONS: ALLOP may increase voluntary EtOH consumption in male mice by altering its reinforcing effects. The lack of significant effect on quinine and water consumption suggests that ALLOP does not simply increase consumption of all fluids.     

Preclinical efficacy evaluations of XK-469: dose schedule,route and cross-resistance behavior in tumor bearing mice

XK-469 is advancing to Phase I clinicaltrials. Preclinical studies were carriedout to assist in clinical applications.Dose-schedule route testing: Singledose IV treatment with XK-469 producedlethality (LD20 to LD 100) above 142 mg/kg.Optimum treatment required total dosages of350 to 600 mg/kg. Furthermore, highindividual IV dosages (100 to 142 mg/kg)were poorly tolerated, producingsubstantial weight loss (8 to 18% of bodyweight), poor appearance, and slow recovery(8 to 12 days). A 1-hour infusion ofdosages more than 140 mg/kg, or BIDinjections 6 hrs apart, did not reducelethality. However, lower individualdosages of 40 to 50 mg/kg/injection IV werewell tolerated and could be given daily toreach an optimum total dose with minimaltoxicities. Likewise, 75 mg/kg/injection IVcould be used every other day to reachoptimal treatment. The necropsy profiles ofdeaths from toxic dosages were essentiallyidentical regardless of schedule (deaths 4to 7 days post treatment). The profileswere: paralytic ileus or gastroparesis; GIepithelial damage; and marrow toxicity.Interestingly, the key lethal events wererapidly reversible and simple to overcomewith lower dosages given daily or everyother day. Based on these results, the highdose, Q21day schedule should be avoided inclinical applications. Instead, a splitdose regimen is recommended (e.g., daily,every other day, or twice weekly). XK-469was also well tolerated by the oral route,requiring 35% higher dosages PO to reachthe same efficacy and toxicity as producedIV. Cross-resistance studies: XK-469resistance was produced by optimumtreatments of IV implanted L1210 leukemiaover seven passage generations. Thisleukemia subline (L1210/XK469) had reducedsensitivity to VP-16 (with a 4.0 log killin IV implanted L1210/XK469 compared to an8.0 log kill against IV implanted L1210/0). It also had a reduction in the sensitivityto 5-FU (with a 2.0 log kill in theimplanted L1210/XK469 compared to a 4.0 logkill against IV implanted L1210/0). Otheragents were approximately as active againstthe resistant tumor, including: Ara-C,Gemzar, Cytoxan, BCNU, DTIC, and CisDDPT. No case of collateral sensitivity wasobserved; i.e., no agent was markedly moreactive against the resistant sublineL1210/XK-469 than against the parent tumorin mice.     

GABA(A)-benzodiazepine receptor complex sensitivity in 5-HT(1A) receptor knockout mice on a 129/Sv background

Previous studies in 5-HT(1A) receptor knockout (1AKO) mice on a mixed Swiss Websterx129/Sv (SWx129/Sv) and a pure 129/Sv genetic background suggest a differential gamma-aminobutyric acid (GABA(A))-benzodiazepine receptor complex sensitivity in both strains, independent from the anxious phenotype. To further investigate these discrepancies, various GABA(A)-benzodiazepine receptor ligands were tested in different behavioral paradigms in 1AKO and wild type (WT) mice on a 129/Sv background. 1AKO and WT mice responded comparably to alprazolam, flumazenil, alcohol and pentylenetetrazol as measured in the stress-induced hyperthermia paradigm. In addition, sedative-anesthetic effects of pentobarbital measured via the righting reflex were similar and a selected dose of diazepam exerted similar anxiolytic effects in both genotypes in the elevated plus maze. In conclusion, 1AKO mice on a 129/Sv background have undisturbed GABA(A)-benzodiazepine receptor sensitivity in contrast to those described on a mixed Swiss Websterx129/Sv background. The anxious phenotype of 1AKO mice seems to occur independent of the GABA(A)-benzodiazepine receptor complex functioning.     

Contribution of the alpha1-GABA(A) receptor subtype to the pharmacological actions of benzodiazepine site inverse agonists

A histidine-to-arginine point-mutation at position 101 in the alpha1-subunit of gamma-aminobutyric acid (GABA)(A) receptors has been shown to switch the in vitro efficacy of Ro 15-4513 from inverse agonism to agonism. In order to assess the consequences of this pharmacological switch in vivo, the motor and proconvulsant effects of Ro 15-4513 were analyzed in knock-in mice containing point-mutated alpha1(H101R)-GABA(A) receptors. Furthermore the influence of the alpha1(H101R) substitution on the efficacy of the beta-carboline inverse agonist DMCM was examined both in vitro and in vivo. Ro 15-4513 (10 mg/kg) increased baseline locomotion and potentiated the convulsant effect of pentylenetetrazole in wild type mice. In alpha1(H101R) mice, Ro 15-4513 decreased locomotion and, at a higher dose (30 mg/kg) it displayed an anticonvulsant action. In vitro, DMCM acted as an inverse agonist at recombinant alpha1beta2gamma2 receptors whereas it potentiated GABA-evoked chloride currents at alpha1(H101R)beta2gamma2 receptors. DMCM was inactive as a convulsant in alpha1(H101R) mice. In keeping with the major contribution of these receptors to the sedative and anticonvulsant properties of benzodiazepine site agonists, the present findings identify the alpha1-GABA(A) receptors as the molecular targets for the allosteric modulation by benzodiazepine site ligands in either direction with regard to the behavioral outputs, sedation/motor stimulation and anticonvulsion/proconvulsion.     

视黄酸在体内外对胃癌细胞转移及其相关蛋白的影响

目的:从体内外探讨全反式视黄酸(ATRA)对人胃癌细胞转移及其相关蛋白影响.方法:胃癌细胞接种到裸鼠脾包膜,每隔两天灌胃给予ATRA 0.7 mg/kg,6周后处死裸鼠,取出所有在脾和肝形成的肿瘤,一些肿瘤被固定和包埋,另一些肿瘤保存在液氮中用于后续实验.用蛋白质印迹法测定蛋白水平;通过免疫组化显示微血管;采用粘附实验测定细胞粘附能力.结果:ATRA灌胃后,脾移植瘤和肝转移瘤受到明显抑制(50％),血管生成也受到抑制.尽管ATRA在体内外调节nm23和mts1/p16蛋白的方式不同,但高比值的nm23:mtsl/p16与低粘附性相关.ATRA在体内外诱导ICAM-1蛋白表达.结论:ATRA显著抑制移植瘤生长及其向肝转移,这一过程可能与对转移相关蛋白nm23,mstl/p16和ICAM-1的调控有关.     

重组L—门冬酰胺酶与其野生型在体外和体内的抗肿瘤作用比较

目的:研究重组L-门冬酰胺酶对肿瘤细胞体外生长及实验性肿瘤的抑制作用.方法:体外培养肿瘤细胞(K562、L1210和P815),分别用倒置显微镜和透射电镜观察肿瘤细胞的形态学,MTT比色法观察细胞增殖率及抑制率,流式细胞仪分析DNA含量.同时根据移植性肿瘤研究方法小鼠接种L1210、P388、Heps及S_(180)瘤株,观察给药后小鼠存活天数和瘤重.结果:体外细胞培养实验证明,重组L-门冬酰胺酶对K562、L1210和P815细胞生长具有显著抑制作用(P<0.01).体内实验表明,重组L-门冬酰胺酶ip能显著延长移植肿瘤小鼠(L1210和P388)的存活天数(P<0.01),并对小鼠移植性肝癌实体瘤(Heps)生长有明显的抑制作用(P<0.01)。重组L-门冬酰胺酶iv对小鼠Heps和S_(180)肿瘤生长有明显的抑制作用(P<0.01).结论:重组L-门冬酰胺酶对肿瘤细胞体外生长及受试肿瘤有明显抑制作用.本结果对重组L-门冬酰胺酶的临床应用提供实验依据.     

Brain-derived neurotrophic factor regulates expression of vasoactive intestinal polypeptide in retinal amacrine cells

Brain-derived neurotrophic-factor (BDNF) is expressed in the retina and controls the development of subtypes of amacrine cells. In the present study we investigated the effects of BDNF on amacrine cells expressing vasoactive intestinal polypeptide (VIP). Rats received three intraocular injections of BDNF on postnatal days (P) 16, 18, and 20. The animals were sacrificed on P22, P40, P60, P80, and P120, and VIP expression in their retinas was detected by immunohistochemistry (P22, P40) and by radioimmunoassay (RIA; P22, P40, P60, P80, P120) to assess the time course of BDNF effects on VIP. A significant increase in the density of VIP-positive amacrine cells was detected in BDNF-treated retinas, and VIP concentration was up-regulated by 150% both at P22 and at P40 with respect to untreated controls. VIP concentration then slowly declined in the treated retinas over a period of 3 months; however, a statistically significant increase of 50% was still detectable on P120. The impact of endogenous BDNF on the regulation of VIP expression in the retina was analyzed in mice homozygous for a targeted deletion of the BDNF gene locus (bdnf-/-). VIP immunohistochemistry revealed a marked reduction of VIP-positive amacrine cells and of VIP-immunopositive processes in the inner plexiform layer of the BDNF knockout mice. Mice lacking BDNF expressed only 5% of the VIP protein in their retinas compared with the retinas of wild-type mice as measured by RIA. Our data show that BDNF is a major regulator of VIP expression in retinal amacrine cells and exerts long-lasting effects on VIP content.