Radix Puerariae modulates glutamatergic synaptic architecture and potentiates functional synaptic plasticity in primary hippocampal neurons

Abstract

Ethnopharmacological relevance: Neurologic disorders are frequently characterized by synaptic pathology, including abnormal density and morphology of dendritic spines, synapse loss, and aberrant synaptic signaling and plasticity. Therefore, to promote and/or protect synapses by the use of natural molecules capable of modulating neurodevelopmental events, such as, spinogenesis and synaptic plasticity, could offer a preventive and curative strategy for nervous disorders associated with synaptic pathology. Radix Puerariae, the root of Pueraria monatana var. lobata (Willd.) Sanjappa&Pradeep, is a Chinese ethnomedicine, traditionally used for the treatment of memory-related nervous disorders including Alzheimer's disease. In the previous study, we showed that the ethanolic extracts of Radix Puerariae (RPE) and its prime constituent, puerarin induced neuritogenesis and synapse formation in cultured hippocampal neurons, and thus could improve memory functions.

Aims of the study: In the present study, we specifically investigated the abilities of RPE and puerarin to improve memory-related brain disorders through modulating synaptic maturation and functional potentiation.

Materials and methods: Rat embryonic (E19) brain neurons were cultured in the absence or presence of RPE or puerarin. At predetermined times, cells were live-stained with DiO or fixed and immunostained to visualize neuronal morphologies, or lysed for protein harvesting. Morphometric analyses of dendritic spines and synaptogenesis were performed using Image J software. Functional pre- and postsynaptic plasticity was measured by FM1-43 staining and whole-cell patch clamping, respectively. RPE or puerarin-mediated changes in actin-related protein 2 were assessed by Western blotting. Neuronal survivals were measured using propidium iodide exclusion assay.

Results: RPE and puerarin both: (1) promoted a significant increase in the numbers, and maturation, of dendritic spines; (2) modulated the formation of glutamatergic synapses; (3) potentiated synaptic transmission by increasing the sizes of reserve vesicle pools at presynaptic terminals; (4) enhanced NMDA receptor-mediated postsynaptic currents, and (5) increased cell viability against naturally occurring cell death. Moreover, upregulation of actin-related protein 2 (ARP2) in RPE and puerarin treated brain neurons suggest that RPE and puerarin induced synaptic plasticity might be associated, at least in part, with ARP2-mediated actin-dependent regulation of spinogenesis.

Conclusions: Our findings indicate that RPE and puerarin might play a substantial role in the morphological and functional maturation of brain neurons and suggest that RPE and puerarin are potentially valuable preventative therapeutics for memory-related nervous disorders.

Keywords: Actin-related protein 2; Dendritic spines; FM1-43; Functional synaptic plasticity; NMDA currents; Puerarin; Radix Puerariae.