Kim Youngkyu, Cho Minju, Paulson Bjorn, Kim Sung-Hoon, Kim Jun Ki
Biomedical Engineering Research Center, Asan Medical Center, Seoul 05505, Republic of Korea.
Department of Anesthesiology and Pain Medicine, Asan Medical Center, University of Ulsan College of Medicine, 88, Olympic-ro 43-Gil, Songpa-gu, Seoul 05505, Republic of Korea.
Microsc Microanal. 2022 May 23:1-8. doi: 10.1017/S1431927622000708.
Among intravital imaging instruments, the intravital two-photon fluorescence excitation microscope has the advantage of enabling real-time 3D fluorescence imaging deep into cells and tissues, with reduced photobleaching and photodamage compared with conventional intravital confocal microscopes. However, excessive motion of organs due to involuntary movement such as breathing may result in out-of-focus images and severe fluorescence intensity fluctuations, which hinder meaningful imaging and analysis. The clinically approved alpha-2 adrenergic receptor agonist dexmedetomidine was administered to mice during two-photon fluorescence intravital imaging to alleviate this problem. As dexmedetomidine blocks the release of the neurotransmitter norepinephrine, pain is suppressed, blood pressure is reduced, and a sedation effect is observed. By tracking the quality of focus and stability of detected fluorescence in two-photon fluorescence images of fluorescein isothiocyanate-sensitized liver vasculature in vivo, we demonstrated that intravascular dexmedetomidine can reduce fluorescence fluctuations caused by respiration on a timescale of minutes in mice, improving image quality and resolution. The results indicate that short-term dexmedetomidine treatment is suitable for reducing involuntary motion in preclinical intravital imaging studies. This method may be applicable to other animal models.
在活体成像仪器中，活体双光子荧光激发显微镜具有能够对细胞和组织进行深入的实时三维荧光成像的优势，与传统的活体共聚焦显微镜相比，其光漂白和光损伤减少。然而，由于呼吸等非自主运动导致的器官过度移动可能会导致图像失焦和严重的荧光强度波动，这会妨碍有意义的成像和分析。在双光子荧光活体成像过程中，给小鼠注射临床批准的α-2肾上腺素能受体激动剂右美托咪定以缓解这一问题。由于右美托咪定可阻断神经递质去甲肾上腺素的释放，从而抑制疼痛、降低血压并产生镇静作用。通过在体内追踪异硫氰酸荧光素敏化的肝脏血管系统的双光子荧光图像中的聚焦质量和检测到的荧光稳定性，我们证明血管内注射右美托咪定可在数分钟的时间尺度上减少小鼠呼吸引起的荧光波动，提高图像质量和分辨率。结果表明，短期右美托咪定治疗适用于减少临床前活体成像研究中的非自主运动。该方法可能适用于其他动物模型。
