Department of Medical Pharmacology, CNR Institute of Neuroscience, Università di Milano, I-20129 Milan, Italy.
J Neurosci. 2012 Feb 8;32(6):1989-2001. doi: 10.1523/JNEUROSCI.2720-11.2012.
Inhibitory (GABAergic) interneurons entrain assemblies of excitatory principal neurons to orchestrate information processing in the hippocampus. Disrupting the dynamic recruitment as well as the temporally precise activity of interneurons in hippocampal circuitries can manifest in epileptiform seizures, and impact specific behavioral traits. Despite the importance of GABAergic interneurons during information encoding in the brain, experimental tools to selectively manipulate GABAergic neurotransmission are limited. Here, we report the selective elimination of GABAergic interneurons by a ribosome inactivation approach through delivery of saporin-conjugated anti-vesicular GABA transporter antibodies (SAVAs) in vitro as well as in the mouse and rat hippocampus in vivo. We demonstrate the selective loss of GABAergic--but not glutamatergic--synapses, reduced GABA release, and a shift in excitation/inhibition balance in mixed cultures of hippocampal neurons exposed to SAVAs. We also show the focal and indiscriminate loss of calbindin(+), calretinin(+), parvalbumin/system A transporter 1(+), somatostatin(+), vesicular glutamate transporter 3 (VGLUT3)/cholecystokinin/CB(1) cannabinoid receptor(+) and neuropeptide Y(+) local-circuit interneurons upon SAVA microlesions to the CA1 subfield of the rodent hippocampus, with interneuron debris phagocytosed by infiltrating microglia. SAVA microlesions did not affect VGLUT1(+) excitatory afferents. Yet SAVA-induced rearrangement of the hippocampal circuitry triggered network hyperexcitability associated with the progressive loss of CA1 pyramidal cells and the dispersion of dentate granule cells. Overall, our data identify SAVAs as an effective tool to eliminate GABAergic neurons from neuronal circuits underpinning high-order behaviors and cognition, and whose manipulation can recapitulate pathogenic cascades of epilepsy and other neuropsychiatric illnesses.
抑制性(GABA 能)中间神经元使兴奋性主神经元集合,以协调海马体中的信息处理。破坏海马体电路中中间神经元的动态募集以及时间上精确的活动,可能表现为癫痫发作,并影响特定的行为特征。尽管 GABA 能中间神经元在大脑中的信息编码中很重要,但用于选择性操纵 GABA 能神经传递的实验工具是有限的。在这里,我们报告了通过递送 Saporin 缀合的抗囊泡 GABA 转运体抗体(SAVAs)在体外以及在体内的小鼠和大鼠海马体中,通过核糖体失活方法选择性消除 GABA 能中间神经元。我们证明了在暴露于 SAVAs 的海马神经元混合培养物中 GABA 能突触而非谷氨酸能突触的选择性丧失、GABA 释放减少以及兴奋/抑制平衡的转变。我们还显示了在 SAVA 微损伤到啮齿动物海马体 CA1 亚区时,囊泡谷氨酸转运体 3 (VGLUT3)/胆囊收缩素/CB(1) 大麻素受体 (+) 和神经肽 Y(+) 局部回路中间神经元的焦点和无差别丧失,中间神经元碎片被浸润的小胶质细胞吞噬。SAVA 微损伤不会影响 VGLUT1(+)兴奋性传入。然而,SAVA 诱导的海马体电路重排引发了网络超兴奋性,与 CA1 锥体神经元的进行性丧失和齿状回颗粒细胞的分散有关。总的来说,我们的数据表明 SAVAs 是一种有效的工具,可以从支持高级行为和认知的神经元电路中消除 GABA 能神经元,并且其操作可以再现癫痫和其他神经精神疾病的致病级联反应。