Department of Veterinary Integrative Biosciences, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843-4458, USA.
Neurotox Res. 2011 Jan;19(1):123-7. doi: 10.1007/s12640-009-9147-5. Epub 2009 Dec 31.
Maintaining calcium ion (Ca²+) homeostasis is crucial for normal neuronal function. Altered Ca²+ homeostasis interferes with Ca²+ signaling processes and affects neuronal survival. In this study, we used homozygous leaner and tottering mutant mice, which carry autosomal recessive mutations in the gene coding for the α(1A) pore forming subunit of Ca(V)2.1 (P/Q-type) voltage-gated calcium channels (VGCC). Leaner mice show severe ataxia and epilepsy, while tottering mice are less severely affected. Leaner cerebellar granule cells (CGC) show extensive apoptotic cell death that peaks at postnatal (P) day 20 and continues into adulthood. Intracellular Ca²+ (Ca²+) concentrations in leaner and tottering mouse Purkinje cells have been described, but Ca²+ concentrations have not been reported for granule cells, the largest neuronal population of the cerebellum. Using the ratiometric dye, Fura-2 AM, we investigated the role of Ca²+ homeostasis in CGC death during postnatal development by demonstrating basal Ca²+, depolarization induced Ca²+ transients, and Ca²+ transients after completely blocking Ca(V)2.1 VGCC. From P20 onward, basal Ca²+ levels in leaner CGC were significantly lower compared to age-matched wild-type CGC. We also compared basal Ca²+ levels in leaner and wild-type CGC to basal Ca²+ in tottering CGC. Potassium chloride induced depolarization revealed no significant difference in Ca²+ transients between leaner and wild-type CGC, indicating that even though leaner CGC have dysfunctional P/Q-type VGCC, Ca²+ transients after depolarization are the same. This suggests that other VGCC are compensating for the dysfunctional P/Q channels. This finding was further confirmed by completely blocking Ca(V)2.1 VGCC using ω-Agatoxin IV-A.
维持钙离子（Ca²+）稳态对于正常神经元功能至关重要。Ca²+稳态的改变干扰了 Ca²+信号转导过程，影响了神经元的存活。在这项研究中，我们使用纯合 leaner 和 tottering 突变小鼠，这些小鼠携带编码钙通道（VGCC）α（1A）孔形成亚基的基因的常染色体隐性突变。leaner 小鼠表现出严重的共济失调和癫痫，而 tottering 小鼠的影响则较轻。leaner 小脑颗粒细胞（CGC）表现出广泛的凋亡性细胞死亡，在出生后（P）第 20 天达到高峰，并持续到成年。已经描述了 leaner 和 tottering 小鼠浦肯野细胞中的细胞内 Ca²+（[Ca²+]（i））浓度，但尚未报道小脑颗粒细胞中的 [Ca²+]（i）浓度，小脑颗粒细胞是小脑最大的神经元群体。使用比率染料 Fura-2 AM，我们通过证明基础 [Ca²+]（i）、去极化诱导的 Ca²+瞬变以及完全阻断 Ca²+（V）2.1 VGCC 后的 Ca²+瞬变，研究了 Ca²+稳态在出生后发育过程中对 CGC 死亡的作用。从 P20 开始，与年龄匹配的野生型 CGC 相比，leaner CGC 的基础 [Ca²+]（i）水平明显较低。我们还将 leaner 和野生型 CGC 的基础 [Ca²+]（i）水平与 tottering CGC 的基础 [Ca²+]（i）水平进行了比较。氯化钾诱导的去极化显示 leaner 和野生型 CGC 之间的 Ca²+瞬变没有显著差异，这表明尽管 leaner CGC 具有功能失调的 P/Q 型 VGCC，但去极化后的 Ca²+瞬变是相同的。这表明其他 VGCC 正在补偿功能失调的 P/Q 通道。使用 ω-Agatoxin IV-A 完全阻断 Ca²+（V）2.1 VGCC 进一步证实了这一发现。
