Department of Developmental Biology, Washington University School of Medicine, St. Louis, MO, USA.
Cardiovascular Division, Department of Medicine, Washington University School of Medicine, St. Louis, MO, USA.
J Gen Physiol. 2023 Sep 4;155(9). doi: 10.1085/jgp.202213310. Epub 2023 Jul 26.
Considerable evidence suggests that day-night rhythms in the functional expression of subthreshold potassium (K+) channels regulate daily oscillations in the spontaneous firing rates of neurons in the suprachiasmatic nucleus (SCN), the master circadian pacemaker in mammals. The K+ conductance(s) driving these daily rhythms in the repetitive firing rates of SCN neurons, however, have not been identified. To test the hypothesis that subthreshold Kv12.1/Kv12.2-encoded K+ channels play a role, we obtained current-clamp recordings from SCN neurons in slices prepared from adult mice harboring targeted disruptions in the Kcnh8 (Kv12.1-/-) or Kcnh3 (Kv12.2-/-) locus. We found that mean nighttime repetitive firing rates were higher in Kv12.1-/- and Kv12.2-/- than in wild type (WT), SCN neurons. In marked contrast, mean daytime repetitive firing rates were similar in Kv12.1-/-, Kv12.2-/-, and WT SCN neurons, and the day-night difference in mean repetitive firing rates, a hallmark feature of WT SCN neurons, was eliminated in Kv12.1-/- and Kv12.2-/- SCN neurons. Similar results were obtained with in vivo shRNA-mediated acute knockdown of Kv12.1 or Kv12.2 in adult SCN neurons. Voltage-clamp experiments revealed that Kv12-encoded current densities in WT SCN neurons are higher at night than during the day. In addition, the pharmacological block of Kv12-encoded currents increased the mean repetitive firing rate of nighttime, but not daytime, in WT SCN neurons. Dynamic clamp-mediated subtraction of modeled Kv12-encoded currents also selectively increased the mean repetitive firing rates of nighttime WT SCN neurons. Despite the elimination of the nighttime decrease in the mean repetitive firing rates of SCN neurons, however, locomotor (wheel-running) activity remained rhythmic in Kv12.1-/-, Kv12.2-/-, and Kv12.1-targeted shRNA-expressing, and Kv12.2-targeted shRNA-expressing animals.
大量证据表明,亚阈值钾 (K+) 通道功能表达的昼夜节律调节了哺乳动物主生物钟视交叉上核 (SCN) 神经元自发放电率的每日波动。然而,驱动 SCN 神经元重复放电率的这些昼夜节律的 K+电导尚未确定。为了检验亚阈值 Kv12.1/Kv12.2 编码 K+通道发挥作用的假设,我们从成年小鼠 SCN 神经元的切片中获得了电流钳记录,这些小鼠在 Kcnh8 (Kv12.1-/-) 或 Kcnh3 (Kv12.2-/-) 基因座中存在靶向破坏。我们发现,Kv12.1-/- 和 Kv12.2-/- SCN 神经元的夜间平均重复放电率高于野生型 (WT) SCN 神经元。相比之下,Kv12.1-/-、Kv12.2-/- 和 WT SCN 神经元的日间平均重复放电率相似,WT SCN 神经元的昼夜差异特征性特征在 Kv12.1-/- 和 Kv12.2-/- SCN 神经元中消除。用体内 shRNA 介导的急性敲低成年 SCN 神经元中的 Kv12.1 或 Kv12.2 也得到了类似的结果。电压钳实验表明,WT SCN 神经元中的 Kv12 编码电流密度在夜间高于白天。此外,Kv12 编码电流的药理学阻断增加了 WT SCN 神经元夜间的平均重复放电率,但不增加白天的平均重复放电率。通过动态钳位介导的建模 Kv12 编码电流的减法也选择性地增加了夜间 WT SCN 神经元的平均重复放电率。然而,尽管 SCN 神经元的平均重复放电率的夜间下降被消除,但运动 (轮跑) 活动在 Kv12.1-/-、Kv12.2-/-和 Kv12.1 靶向 shRNA 表达以及 Kv12.2 靶向 shRNA 表达的动物中仍然保持节律性。