Fig. 1: briefCO₂ stimulation generates early and delayed responses.

a Head-fixed sedated mice, implanted with a chronic cranial window over the barrel cortex, were exposed to a brief and strong hypercapnic stimulation (briefCO₂: 20%, 10 s) or to whole pad whisker stimulation (5 Hz, 5 s). Respiration was monitored with a thermocouple placed in front of the nostril and blood flow imaged with two-photon imaging. b Top, CO₂ concentration measured at the nostril reached a plateau within 2 s. Middle and inset, detection of temperature peaks enabled the respiratory rate to be computed (bottom, single trial), which increased after a delay of about 2 s. c Top left, a pial arteriole labeled with Texas Red injected i.v. The diameter was measured with line-scan acquisitions drawn perpendicular to the vessel (white dotted line). Top right, Representative example of a pial arteriole dilation upon briefCO₂. Bottom left, Average responses to briefCO_2. Diameter changes are expressed in z score to minimize the contribution of spontaneous 0.1 Hz fluctuations. Dilation (pink trace) lagged the hyperventilation (blue trace) by ~ 4 s (see the arrows, paired data, n = 16 arterioles, 12 mice, 2–5 stimulations per vessel). Bottom right, quantification of the onsets (two-sided Wilcoxon rank sum test, ****p < 0.0001). d Top, the diameter of penetrating arterioles was calculated from lumen area. Bottom left and right—as for pial arterioles, hyperventilation (blue trace) precedes dilation (pink trace) by ~ 4 s (paired data, n = 13 arterioles, 11 mice, 2–5 stimulations per vessel, two-sided Wilcoxon rank sum test, ***p = 0.0002). The arrows indicate the onset time to reach 10% of the fit of the curves. Data are represented as mean ± SD (shadings).