Fig. 4. Low toxic Pd(bpy) derivatives for neuronal studies.

a Chemical structures of the Pd(bpy) derivatives. b Evaluation of the effect of Pd(bpy) for NGF-induced neurite outgrowth of PC12 cells. Left: representative fluorescent images of PC12 cells treated with or without Pd(bpy). PC12 cells were visualized using Calcein-AM. Right: concentration-dependency of Pd(bpy) for neurite outgrowth in the PC12 cells after treatment of 1 ng/mL NGF. (n = 20). Scale bars, 50 μm. c Concentration-dependency of Pd(bpy) derivatives on the cell growth in undifferentiated PC12 cells. (n = 3). d The abnormal Ca²⁺ response by Pd(bpy) (black) or Pd(sulfo-bpy) (red) in the cultured cortical neurons. Left: representative trace of the Ca²⁺ response induced by 30 µM Pd(bpy) (black) or Pd(sulfo-bpy) (red). Right: concentration-dependence curves for Pd(bpy) (black) or Pd(sulfo-bpy) (red). (n = 30). e Ca²⁺ responses of Pd(bpy) derivatives for mGlu1(N264H) mutant in HEK293 cells. Left: averaged Δratio induced by 3 µM Pd(bpy) derivatives. (n = 15 for Pd(EG-bpy) and Pd(di-sulfo-bpy). n = 20 for Pd(bpy), Pd(OH-bpy), and Pd(sulfo-bpy)). Data are presented as mean ± s.e.m. f Concentration-dependent curves for Pd(sulfo-bpy) (red) and Pd(OH-bpy) (green) in HEK293 cells expressing the mGlu1(N264H) mutant (n = 20). g Solubility assay of the Pd(bpy) (black) and Pd(sulfo-bpy) (red) in ACSF using UV-vis spectroscopy. Linear increment of absorbance at 312 nm corresponding to the palladium complexes indicates the solubility of the complexes. The concentration dependency of the absorbance of Pd complexes in the high or low concentration range is shown in the left or right, respectively (n = 3). Data are presented as mean ± s.e.m.