Fig. 2: Exogenous CO₂ prevents the development of the neurovascular response in the somatosensory cortex in rats.

a Schematic depiction of the neurovascular unit illustrating the central hypothesis and the experimental approach taken to study the potential role of CO₂ as a signaling molecule that mediates the neurovascular coupling response. A surplus of exogenous CO₂ was given in the inspired air (hypercapnia) in order to saturate the brain CO₂-sensitive vasodilatory mechanism. If the hypothesis is correct, then in the presence of surplus of exogenous CO₂, any extra metabolic CO₂ generated as a result of increased neuronal activity should have no additional effect on local cerebral blood flow (CBF) and the neurovascular response should be blocked. If the mechanisms other than CO₂ signaling mediate the neurovascular coupling response, then the response should not be affected by the excess of exogenous CO₂. b CBF and blood oxygen level dependent (BOLD) signals in the forepaw region of the somatosensory cortex (S1FL) in anesthetized rats were recorded using an arterial spin labeling (ASL) sequence with T2* weighted imaging. Somatosensory pathways were activated by electrical stimulation of the forepaw. RF radiofrequency. c CBF and BOLD responses in the S1FL region induced by electrical forepaw stimulation (3 Hz, 1.5 mA) at baseline, in conditions of 5% and 10% inspired CO₂, after the administration of carbonic anhydrase inhibitor acetazolamide (10 mg kg⁻¹, i.v.), and in conditions of 5% and 10% inspired CO₂, applied concomitantly with systemic carbonic anhydrase inhibition with acetazolamide. Data are presented as mean values ± SEM (shaded areas denote error bands). Insets show representative activation maps illustrating mean BOLD signal changes in response to somatosensory stimulation. Color bar: t-score from statistical paramagnetic mapping mixed-effects analysis, p < 0.05 (uncorrected). d Representative ASL images illustrating CBF at rest (normocapnia) and in conditions of 10% inspired CO₂. Overlaid (false color scale) illustrates CBF response in the S1FL region induced by forepaw stimulation. e, f Summary data illustrating the effect of 5% and 10% inspired CO₂, given before and after systemic administration of acetazolamide, on CBF and BOLD responses recoded in the S1FL region of the cortex. P values, mixed-model ANOVA followed by Holm–Sidak multiple comparison test. g Summary data illustrating peak changes in brain tissue pH (S1FL cortical region) following administration of acetazolamide, as well as in response to 5% and 10% inspired CO₂, given before and after systemic administration of acetazolamide. P values, Kruskal–Wallis test followed by Dunn’s multiple comparison test. In the box-and-whisker plots the central dot indicates the mean, the central line indicates the median, the box limits indicate the upper and lower quartiles, and the whiskers show the minimum–maximum range of the data. Numbers in parentheses indicate sample sizes (number of animals per experimental group). Source data are provided as a Source Data file.