PMC

Precapillary sphincters are the key gatekeepers of capillary blood flow. (A) Mathematical modeling of flow distribution in a reconstructed cerebrovascular tree containing one PA and venule (), under resting conditions. (B) Summary of calculated average blood flows among all PA, precapillary sphincter (Sphinc), and first- to third-order capillary segments in the network before (Rest) and during WP stimulation. The groups labeled with “wo” denote calculations of WP stimulations in which either the PA, precapillary sphincter, and first- to third-order capillary segments were disallowed from dilating. Note the precapillary sphincter has the strongest influence on network flows, as shown in the dashed box. (C) Summary of calculated blood flows in the vessel segments before (Rest) and during ET1 puff. Similar to B, calculations of the puff where segments were disallowed from constricting are shown in the “wo”-labeled groups. A.U., arbitrary units. (D) Mathematical modeling of pressure distribution in the resting state. Inlet pressure into the PA was set to 25 mmHg, whereas all outlet pressures were set to 5 mmHg. (E) Summary of the average pressures in PA, precapillary sphincter, and first- to third-order capillaries in the network normalized to the average pressure in the PA. (F) Normalized pressure change at PA, precapillary sphincter, and first- to third-order capillaries by dilation or constriction. (G) Representative image of mural cells (red) and αSMA (green) staining among the PA (red inset), precapillary sphincter (magenta inset), first-order capillaries (blue inset), second-order capillaries (green insert), and third-order capillaries (orange inset); see insets for enlargements. (H) Summarized intensity of αSMA in pericytes on precapillary sphincter and first-, second-, or third-order capillaries normalized to the intensity from αSMA intensity on the PA. Note that αSMA is significantly more expressed in precapillary sphincters than PA and first- to third-order capillaries, whereas all first- to third-order capillaries contained less αSMA than PA. Individual vessel segments are compared to all other vessel segments, and the highest P value is illustrated. N = 3, n = 31. N denotes total number of animals; n denotes total number of vessels. Linear mixed effect models were used, followed by Tukey post hoc tests. Data shown as mean ± SEM. (I) Relationship between normalized pressure and normalized αSMA density at peak dilation upon WP stimulation. * indicates P < 0.05, ** indicates P < 0.001, *** indicates P < 0.0001.

a Left panel: Maximal intensity projected in vivo two-photon laser scanning microscopy image of an NG2-dsRed mouse barrel cortex. An indentation of the capillary lumen is observed at the branching of the PA and is encircled by bright dsRed cell(s) (dashed insert). This structure is denoted as a precapillary sphincter. Immediately after the sphincter, a sparsely dsRed-labeled distention of the capillary lumen is observed, which we refer to as the bulb. Right panels: Single z-plane showing overlay, FITC-channel, and dsRed channel of the dashed insert. Arrows indicate the PA (red), sphincter (blue), bulb (green), and 1^st order capillary (yellow). b–d Local TPLSM projections of precapillary sphincters in the cortex of a thinned skull mouse in vivo (b), an awake mouse harboring a chronic cranial window in vivo (c) with white arrows marking the precapillary sphincter, and an ex vivo coronal slice of a FITC-conjugated lectin (green) stained NG2-dsRed mouse (red) with DAPI-stained (blue) nuclei (d). The precapillary sphincter cell nucleus is arched, as it follows the cell shape, and is marked by a white arrowhead. e Schematic of a PA with the a a precapillary sphincter at the proximal branch point. The illustration is based on confocal imaging of coronal slices ex vivo and the exact morphology and location of NG2-dsRed positive cells and their DAPI stained nuclei are shown. For the complete figure including a venule, see Supplementary Fig. .

a An xz-projection of a z-stack covering an entire PA (left) was reconstructed (middle) and converted into a computational model of a PA and associated first and second order capillaries. Model simulation (right) including the sphincters (green, marked by asterisks), or without the sphincters (red), observed in the two proximal branches along the PA gave rise to higly divergent pressure profiles along the first order capillaries (highlighted boxes in the model without sphincters). b Pressure drop across the sphincter depends on its diameter. Focusing on the first sphincter (left panel), the pressure drop across the sphincter (ΔP_Sphincter) relative to the pressure at the branch point (P_PA) was calculated as a function of sphincter diameters (x∙D_Sphincter) under resting conditions (middle) or upon functional stimulation (right, using the relative changes in dilation from Fig. ) with the sphincter (green curves) or without (red curves, where diameter of the sphincter is equal to the first order capillary). c The degree of sphincter contraction correlates to the pressure in the PA. With increasing inlet pressures into the PA (20 mmHg: blue, 25 mmHg: green, and 30 mmHg: red), the PA pressure increases (dashed curves) and the sphincter must contract to maintain a relatively low pressure into the first order capillary (full curves). The difference between the pressures in the PA and the first order capillary is the pressure drop across the sphincter (ΔP_Sphincter, see inset). d Flow reduction and phase separation effects due to the sphincter. The ratios of blood flow (rQ_BF, blue curve) and RBC flow (rQ_RBC, green curve) with and without the sphincter (rQ_{with_Sphincter}/rQ_{without_Sphincter}) was calculated as a function of sphincter diameter. Both flows correlate proportionally with diameter but the RBC flow remains lower due to plasma skimming. Source data are provided as a Source Data file.

NO donor SNAP-induced vasodilation is mediated by cGMP in capillary pericytes. (A) An average-projected image from a local two-photon image stack with PA, precapillary sphincter (Sphinc), and first- to third-order capillaries. (Scale bar, 10 µm.) (B) Representative traces of SNAP-puff-induced vasodilation before and after topical application of cGMP inhibitor soluble guanylyl cyclase inhibitor (ODQ). (C) Comparison of relative diameter change at PA, precapillary sphincter, and first- to third-order capillaries, induced by SNAP puff before and after ODQ. Before: N = 7, n = 7; after: N = 6, n = 6. N denotes total number of animals; n denotes total number of vessels. (D–F) Comparison of WP stimulation-induced vasodilation at PA, precapillary sphincter, and first- to third-order capillaries before and after ODQ, with (D) representative traces of WP stimulation-induced vasodilation before and after topical application of ODQ, (E) dilation amplitude, and (F) dilation duration of half-peak. Before: N = 7, n = 7; after: N = 6, n = 6. Linear mixed effect models were used, followed by Tukey post hoc tests. Data shown as mean ± SEM. * indicates P < 0.05, ** indicates P < 0.001, *** indicates P < 0.0001.