Fig. 1
From: Regulation of density of functional presynaptic terminals by local energy supply
Functional terminal density in response to 5AP bursts is nonlinearly associated with extracellular Mg2+ concentration. (a) The schematic experimental procedures. Magnesium Green-AM ester (MgGrn) staining and imaging were performed at basal condition (close to natural culturing state, without eliciting any stimulus). After MgGrn imaging, vesicle turnover was detected by FM dye under field stimulations, such as bursting stimulations (e.g. 5AP bursts). For the bursting stimulation, 30 action potentials (APs) were divided into 6 bursts (inter-burst-interval was 10 s), each of which contained 5 APs at 100 Hz. The MgGrn and FM dye imaging procedures were combined (e.g. for the experiment in b) or conducted separately based on different experimental designs. (b) Neuron cultures with [Mg2+]o of 0.8, 1.2 or 2.0 mM in culture medium (for 48 hr to 2 weeks) were marked by MgGrn to reveal their [Mg2+]i level, and then functional terminals were detected by FM4-64 under 5AP bursts (as described in a) at the same area of interests (AOIs). Pseudo-color scale: fluorescent intensity. (c) Bell-shape association between functional terminal density in response to 5AP bursts (N 5AP ) and [Mg2+]o (n = 3–5 coverslips, Gaussian curve fitting, R2 = 0.84). (d) Bell-shape association between [Mg2+]i (MgGrn fluorescence) and [Mg2+]o (n = 3–5 coverslips, Gaussian curve fitting, R2 = 0.86). The mean ± SEM of coverslips was presented. For the measurement of N 5AP and [Mg2+]i see Methods