Timic Stamenic Tamara, Feseha Simon, Fine-Raquet Brier, Tadic Vasilije P, Todorovic Slobodan M
Department of Anesthesiology, University of Colorado, Aurora, CO, United States.
Department of Neuroscience, University of Colorado, Aurora, CO, United States.
Exp Biol Med (Maywood). 2025 May 16;250:10553. doi: 10.3389/ebm.2025.10553. eCollection 2025.
Although substantial progress has been made in the last three decades towards our understanding of how general anesthetics (GAs) act at the molecular level, much less is known about how GAs cause loss of consciousness at the level of neuronal networks. The role of thalamus as an important brain region in anesthetic-induced hypnosis is relatively well established, but the specific roles of voltage-gated ion channels in different functional regions of the thalamus in anesthetic mechanisms are not well studied. To address this gap in knowledge, we selectively silenced the gene that encodes the low-threshold-activated Ca3.1 T-type voltage-gated calcium channel subunit by injecting short-hairpin RNA (shRNA) into midline and intralaminar - nonspecific thalamus (MIT) and sensory - specific ventrobasal (VB) thalamic nuclei in wild-type (WT) mice. Control animals were injected with scrambled shRNA. To validate our silencing approach, we performed patch-clamp experiments in acute thalamic slices . In injected animals we determined anesthetic endpoints such as hypnosis measured with loss of righting reflex (LORR) and immobilization measured with loss of withdrawal reflex (LOWR) after administration of a traditional volatile GA isoflurane. Effective Ca3.1 channel knock-down was documented by greatly diminished amplitudes of T-currents and absence of rebound burst firing in our patch-clamp recordings from thalamic slices. We found that knocking down Ca3.1 channels in MIT significantly decreased inhaled isoflurane concentration that is required to induce LORR, but it did not affect speed of anesthetic induction and the immobilizing effect of isoflurane. In contrast, knocking down the Ca3.1 channel in the VB thalamus did not affect any of the measured anesthetic endpoints. Hence, we concluded that Ca3.1 channels in nonspecific MIT thalamus have a preferential role in anesthetic hypnosis when compared to the sensory VB thalamus.
尽管在过去三十年里，我们在理解全身麻醉药（GAs）如何在分子水平发挥作用方面取得了重大进展，但对于GAs如何在神经网络层面导致意识丧失却知之甚少。丘脑作为麻醉诱导催眠过程中的一个重要脑区，其作用相对已得到较好的确立，但丘脑不同功能区域的电压门控离子通道在麻醉机制中的具体作用尚未得到充分研究。为了填补这一知识空白，我们通过向野生型（WT）小鼠的中线和层内 - 非特异性丘脑（MIT）以及感觉特异性腹侧基底（VB）丘脑核团注射短发夹RNA（shRNA），选择性地沉默了编码低阈值激活的Ca3.1 T型电压门控钙通道亚基的基因。对照动物注射乱序shRNA。为了验证我们的沉默方法，我们在急性丘脑切片上进行了膜片钳实验。在注射动物中，我们在给予传统挥发性麻醉药异氟烷后，确定了麻醉终点，如通过翻正反射消失（LORR）测量的催眠和通过退缩反射消失（LOWR）测量的制动。我们从丘脑切片的膜片钳记录中发现，T电流幅度大幅减小以及没有反弹爆发放电，证明了Ca3.1通道的有效敲低。我们发现，敲低MIT中的Ca3.1通道显著降低了诱导LORR所需的吸入异氟烷浓度，但不影响麻醉诱导速度和异氟烷的制动效果。相比之下，敲低VB丘脑中的Ca3.1通道对任何测量的麻醉终点均无影响。因此，我们得出结论，与感觉性VB丘脑相比，非特异性MIT丘脑中的Ca3.1通道在麻醉催眠中具有优先作用。
