以N-甲基-D-天門冬胺酸受體功能低下小鼠模式評估肌胺酸對思覺失調症相關行為與認知缺損之療效

Abstract

思覺失調症是一種嚴重且耗費社會成本甚鉅的心智疾患。麩胺酸神經傳導系統的N-甲基-D-天門冬胺酸受體功能低下為思覺失調症新興假說，相關的潛力藥物能緩解病人不同面向的症狀，尤其是負向及認知症狀。根據此假說，增強N-甲基-D-天門冬胺酸受體功能可達到治療效果，而治療藥物作用在受體上的甘胺酸調節位，可避免因麩胺酸結合位被過度刺激而引發神經毒性。甘胺酸又為N-甲基-D-天門冬胺酸受體的共同致效劑，N-甲基-D-天門冬胺酸受體附近的甘胺酸濃度由甘胺酸型一轉運體負責調節，並維持其在低飽和狀態。肌胺酸為一種甘胺酸型一轉運體抑制劑，可抑制甘胺酸回收，增加突觸間甘胺酸濃度，進而提升N-甲基-D-天門冬胺酸受體功能。儘管過去已有研究指出肌胺酸能反轉N-甲基-D-天門冬胺酸受體功能低下現象，但有效劑量及療效背後的機制為何仍不清楚。因此，本論文設計了三個研究，試圖探討肌胺酸劑量效果，並以兩種N-甲基-D-天門冬胺酸受體功能低下動物為模型，觀察肌胺酸對思覺失調相關行為與認知缺損的療效。在研究一我們發現，單一注射低至中等劑量（250, 200, 1000及2000 mg/kg）的肌胺酸不影響小鼠自主活動量且未產生明顯毒性反應。在研究二，我們使用一種N-甲基-D-天門冬胺酸受體拮抗劑–MK-801來引發思覺失調症相關行為缺損。實驗結果發現，單一注射肌胺酸不影響MK-801誘發的過度活動與刻板行為（除了最高劑量2000 mg/kg），然而，在1000 mg/kg的劑量下，肌胺酸能達到有效改善由MK-801引發驚跳反應、探索行為與恐懼制約行為缺損的效果。同時，我們透過正子斷層掃描腦造影技術發現，MK-801會明顯引發腦部異常活化現象，而給予肌胺酸則能有效緩解此異常。最後，在研究三，我們以先天性N-甲基-D-天門冬胺酸受體功能低下的基因缺損小鼠(絲氨酸消旋酶基因剔除鼠)為模型，給予肌胺酸治療後，發現基因剔除小鼠在認知行為方面的缺損得到改善。綜合以上，本研究顯示N-甲基-D-天門冬胺酸功能低下的小鼠模型確實能再現思覺失調相關的行為與認知缺損，而給予肌胺酸後可改善這些缺損，支持肌胺酸為有效治療思覺失調症的潛力藥物。

Schizophrenia is a severe mental disorder, which results in tremendous cost to our society. The hypofunction of the N-methyl-d-aspartate (NMDA) glutamate transmitter system is an emerging hypothesis for schizophrenia, especially for negative and cognitive symptoms. This hypothesis has suggested promising pharmacological agents to ameliorate full range of schizophrenia symptoms. Based upon this hypothesis, therapeutic agents need to enhance the activity of NMDA receptor (NMDAR) via glycine modulation site for avoiding toxicity induced by overstimulation of glutamate binding sites. Glycine, a co-agonist at the NMDAR complex, is maintained at a low sub-saturating level in the vicinity of NMDAR by glycine transporter 1 (GlyT-1). To increase glycine level, sarcosine (N-methylglycine), a GlyT-1 inhibitor, can increase glycine concentration within synaptic cleft to facilitate NMDAR function. Despite emerging evidence on sarcosine in restoring NMDA hypofunction, the effective dosage and the mechanism for its therapeutic effect remain much unclear. In this thesis, three studies were designed and conducted to investigate the dosage and effect of sarcosine on the amelioration of behavioral and cognitive deficits in two mouse models of NMDAR hypofunction. In Study 1, we found that a single injection of sarcosine at low to medium doses (i.e., 250, 500 and 1000 mg/kg, i.p.) did not affect spontaneous locomotor activity in mice and no obvious toxicity was found in these mice as well. In Study 2, MK-801, a NMDAR antagonist, was used to induce schizophrenia-like behavior in male mice. A single administration of sarcosine (except the highest dose, 2000 mg/kg) had no effect on MK-801 induced hyperlocomotion and stereotypy. Intriguingly, a single injection of sarcosine at 1000 mg/kg can alleviate MK-801 induced cognitive deficits in the prepulse inhibition, holeboard task and fear conditioning task. In complementary to behavioral improvement, taking advantage of microPET scan with 18F-fluorodeoxyglucose to visualize brain activity, we also found that sarcosine can ameliorate the alteration of brain activity induced by MK-801 in male mice. In Study 3, serine racemase-null mutant (SR-/-) mice were further used as a genetic NMDAR hypofunction mouse model of schizophrenia. A single injection of sarcosine also successfully rescued observed behavioral and cognitive deficits reported in these male SR-/- mice. Collectively, our mouse models of NMDAR hypofunction that recapture some schizophrenia-like behavioral and cognitive deficits in mice can be ameliorated by a single injection of sarcosine. Findings from this research support the therapeutic effect of sarcosine and its potential in the treatment of schizophrenia.