藉由內視鏡在活體內觀察經歷時差時視交叉上核之神經活性

Abstract

地球上的生物依靠體內的晝夜節律以應對二十四小時的日夜週期變化，而在哺乳類動物中，下視丘的視交叉上核(Suprachiasmatic nucleus, SCN)作為中樞節律器調控眾多生理功能。現代社會中人們頻繁地到世界各地旅遊並跨越不同時區，因此常受時差(jet lag)影響產生睡眠障礙。過去對於時差的研究使用活體外腦片紀錄的方式，發現視交叉上核中的神經細胞的時鐘基因在經歷時差後變得較不同步，並需要幾天的時間逐漸重新適應到新的時區。然而究竟在活體內視交叉上核內的神經活性如何被時差影響仍然未知。藉由在梯度折射率透鏡(gradient index lens, GRIN Lens)、鈣離子影像技術及雙光子顯微鏡，我們得以在清醒小鼠內紀錄視交叉上核內的神經活性。我們發現視交叉上核內神經的鈣離子活動相關性會迅速被外界光線提升，且不會在經歷時差後有所降低；另一方面視交叉上核神經的基礎鈣離子濃度原本具有日夜週期的變化，但卻會在經歷時差後被擾亂，且部分神經會重新適應到新時區。而無法重新適應的神經們彼此之間又具有較強的鈣離子活動相關性，因此他們可能是屬於同一未知功能的神經迴路。綜上所述我們的結果顯示鈣離子活動和基礎鈣離子濃度應該是被兩種不同機制調控，其中後者可能是由生理時鐘基因調控，並負責同步行為與外界光週期。

Living creatures on Earth rely on their intrinsic circadian rhythms to anticipate the external day and night cycle. The suprachiasmatic nucleus (SCN) in the mammalian hypothalamus serves as the central pacemaker to coordinate individual periphery oscillators and govern various physiological functions critical to health. When rapidly crossing several time zones by flight, people often experience jet lag and suffer unsettled sleep-wake cycles. A previous study using in vitro single-cell imaging has revealed that the phases of PER2, a core clock gene, expressions in the SCN neurons become less synchronized after jet lag and require a couple of days to gradually be re-entrained to the new light-dark cycle. However, further in vivo evidence about how jet lag affects SCN neuronal activities is still deficient. In this study, by a combination of gradient index (GRIN) lenses, Ca2+ imaging, and two-photon microscopy, we were able to record single-cell SCN neuronal activities in head-fixed awake mice. We found that the synchrony of transient Ca2+ activities of SCN neurons was acutely enhanced by external light and not altered after jet lag exposure; on the other hand, basal Ca2+ levels of most neurons originally exhibiting daily fluctuations higher in the daytime were disturbed by jet lag, and some neurons achieved re-entrainment of their basal Ca2+ fluctuations to the new light-dark cycle. Moreover, those SCN neurons incapable of re-entrainment exhibited stronger correlations of transient Ca2+ activities to each other, implying they might belong to the same circuit with unknown functions instead of photoentrainment. Together, our findings demonstrate that transient Ca2+ activities and basal Ca2+ levels could be controlled by different mechanisms, with the former acutely responding to light input and the latter probably mediated by clock genes and accounting for photoentrainment of behavior rhythms.