Fig. 3: Microglia regulate ATP but not adenosine-induced cerebral blood flow (CBF) increase.

a Schematic of combining CBF measurement (using laser speckle contrast imaging) and nano-injector guided intracisternal magna (ICM) injection of vasoactive chemicals (Created in BioRender. Lab, K. (2024) https://BioRender.com/l61h464). b Representative images of the CBF responses to ICM injection of acetylcholine (Upper, 1 mM, 3 μl) or adropin peptides (Lower, 0.1 mg/ml, 3 μl). Each experiment was repeated independently for five times with similar results. c, d Real-time CBF changes to ICM-injection of acetylcholine (Ach, c) or adropin (d). The bar indicates the 10-minute injection period. e Representative images of the CBF responses to ICM injection of ATP (Upper, 1.5 μl, 10 mM) or adenosine (ADO, Lower, 1.5 μl, 10 mM). f, h Real-time CBF responses to ICM-injection of ATP (f) or adenosine (h). g Does-responses of CBF changes after ICM-injection of ATP. The injection volume was 1.5 μl in all conditions. Each dot represents an individual mouse. Sample sizes are n = 6, 5, 7, and 3 for the 0.5 mM, 5 mM, 10 mM, and 20 mM conditions, respectively. Data are presented as mean ± s.e.m. Source data are provided as a Source Data file. i Does-responses of CBF changes after ICM-injection of adenosine. The injection volume was 1.5 μl in all conditions. Each dot represents an individual mouse. Sample sizes are n = 4, 5, 5, and 6 for the 0.5 mM, 5 mM, 10 mM, and 20 mM conditions, respectively. Data are presented as mean ± s.e.m. Source data are provided as a Source Data file. j Schematics of sequential measurements of CBF responses in the same mouse at the baseline, after PLX3397-treatment, and at microglia repopulation state. k, l Comparison of the CBF responses to ICM-ATP (10 mM, 1.5 μl) (k) or adenosine (l) injection in the same mice at the baseline, PLX-treated, and repopulation state, respectively. Each dot indicates an individual mouse. n = 8 mice for each condition. Data are presented as mean ± s.e.m. p-values were determined by one-way ANOVA for paired samples with Tukey post hoc test. Source data are provided as a Source Data file. m, n CBF peak value to ICM-ATP (10 mM, 1.5 μl) (m) or adenosine (n) injection in the same mice at the baseline, PLX-treated, and repopulation state, respectively. Each dot indicates an individual mouse. n = 8 mice for each condition. Data are presented as mean ± s.e.m. p-values were determined by one-way ANOVA for paired samples with Tukey post hoc test. Source data are provided as a Source Data file. o, p Representative images of the CBF baseline and responses to ICM-injection of ATPγS (10 mM, 1.5 μl) at the time-points i, ii and iii (indicated by arrowheads in p). All ICM-ATPγS-injected mice expired within 20 min (n = 5).