A Novel Striated Muscle-Specific Myosin-Blocking Drug for the Study of Neuromuscular Physiology.
A Novel Striated Muscle-Specific Myosin-Blocking Drug for the Study of Neuromuscular Physiology.
复制标题
DOI：
10.3389/fncel.2016.00276
复制
发表时间：
2016
影响因子：
5.3
通讯作者：
Gould TW
中科院分区：
文献类型：
--
作者：
Heredia DJ;Schubert D;Maligireddy S;Hennig GW;Gould TW
将神经动作电位(AP)传递到肌肉AP的失败被称为神经肌肉传递失败(NTF)。尽管突触功能障碍存在于多种神经肌肉疾病中，神经传递受损也会导致肌肉疲劳，但由于肌肉随后产生的运动，通过测量成功转导的肌肉AP来直接评估神经传递是困难的。此外，用于研究成年神经肌肉接头神经递质释放的电压门控钠通道抑制剂在胚胎组织中无效，使得几乎不可能精确测量胚胎致死小鼠突变体的任何方面的神经传递。在这项研究中，我们利用六六六(3-(N-butylethanimidoyl)-4-hydroxy-2H-chromen-2-one)，以前在骨骼肌肌球蛋白抑制剂的小分子筛查中发现，在不影响膜电流的情况下抑制运动。与这一筛选的先前确定的药物不同，如N-苄基-对甲苯磺酰胺(BTS)，它抑制骨骼肌肌球蛋白ATPase活性，但也阻断神经传递，BHC选择性地阻断神经诱发的肌肉收缩，而不影响神经递质的释放。这一特征使得胚胎和成年小鼠的神经传递得到了详细的描述。在BHC存在的情况下，以高达20赫兹的频率紧张性刺激膈神经产生的神经性AP被成功地传递到肌肉AP。在较高的膈神经刺激率时，可观察到NTF。NTF是间歇性的，以失败的肌肉AP后成功的肌肉AP为特征，成功传输肌肉AP的百分比随着时间的推移而减少。在六六六存在的情况下，未能产生NTF的神经刺激率同样也未能产生峰值肌肉纤维缩短，这是通过一种新的肌肉疲劳光学方法检测的，即在表达遗传编码钙指示剂GCaMP3的整个肌肉细胞群体中检测到的峰值胞浆钙瞬变强度的丧失。最重要的是，BHC首次对胚胎小鼠的突触传递、钙信号和疲劳进行了详细的分析，例如这里报道的Vamp2突变，这些突变在出生前或出生时死亡。总而言之，这些研究说明了六六六在稳定测量神经肌肉功能方面的广泛用途。
The failure to transmit neural action potentials (APs) into muscle APs is referred to as neuromuscular transmission failure (NTF). Although synaptic dysfunction occurs in a variety of neuromuscular diseases and impaired neurotransmission contributes to muscle fatigue, direct evaluation of neurotransmission by measurement of successfully transduced muscle APs is difficult due to the subsequent movements produced by muscle. Moreover, the voltage-gated sodium channel inhibitor used to study neurotransmitter release at the adult neuromuscular junction is ineffective in embryonic tissue, making it nearly impossible to precisely measure any aspect of neurotransmission in embryonic lethal mouse mutants. In this study we utilized 3-(N-butylethanimidoyl)-4-hydroxy-2H-chromen-2-one (BHC), previously identified in a small-molecule screen of skeletal muscle myosin inhibitors, to suppress movements without affecting membrane currents. In contrast to previously characterized drugs from this screen such as N-benzyl-p-toluene sulphonamide (BTS), which inhibit skeletal muscle myosin ATPase activity but also block neurotransmission, BHC selectively blocked nerve-evoked muscle contraction without affecting neurotransmitter release. This feature allowed a detailed characterization of neurotransmission in both embryonic and adult mice. In the presence of BHC, neural APs produced by tonic stimulation of the phrenic nerve at rates up to 20 Hz were successfully transmitted into muscle APs. At higher rates of phrenic nerve stimulation, NTF was observed. NTF was intermittent and characterized by successful muscle APs following failed ones, with the percentage of successfully transmitted muscle APs diminishing over time. Nerve stimulation rates that failed to produce NTF in the presence of BHC similarly failed to produce a loss of peak muscle fiber shortening, which was examined using a novel optical method of muscle fatigue, or a loss of peak cytosolic calcium transient intensity, examined in whole populations of muscle cells expressing the genetically-encoded calcium indicator GCaMP3. Most importantly, BHC allowed for the first time a detailed analysis of synaptic transmission, calcium signaling and fatigue in embryonic mice, such as in Vamp2 mutants reported here, that die before or at birth. Together, these studies illustrate the wide utility of BHC in allowing stable measurements of neuromuscular function.
登录
查看更多内容
DOI：
10.1083/jcb.81.2.275
发表时间：
1979-05
期刊：
The Journal of cell biology
影响因子：
--
作者：
Heuser JE;Reese TS;Dennis MJ;Jan Y;Jan L;Evans L
通讯作者：
Evans L
影响因子：
3.7
作者：
Bloemberg D;Quadrilatero J
通讯作者：
Quadrilatero J
影响因子：
3.3
作者：
BAZZY, AR;DONNELLY, DF
通讯作者：
DONNELLY, DF
影响因子：
5.5
作者：
Hennig, GW;Smith, CB;Smith, TK
通讯作者：
Smith, TK
影响因子：
3.1
作者：
Fajardo VA;Smith IC;Bombardier E;Chambers PJ;Quadrilatero J;Tupling AR
通讯作者：
Tupling AR