Ultra-Slow Single-Vessel BOLD and CBV-Based fMRI Spatiotemporal Dynamics and Their Correlation with Neuronal Intracellular Calcium Signals

Abstract

Functional MRI has been used to map brain activity and functional connectivity based on the strength and temporal coherence of neurovascular-coupled hemodynamic signals. Here, single-vessel fMRI reveals vessel-specific correlation patterns in both rodents and humans. In anesthetized rats, fluctuations in the vessel-specific fMRI signal are correlated with the intracellular calcium signal measured in neighboring neurons. Further, the blood-oxygen-level-dependent (BOLD) signal from individual venules and the cerebral-blood-volume signal from individual arterioles show correlations at ultra-slow (<0.1 Hz), anesthetic-modulated rhythms. These data support a model that links neuronal activity to intrinsic oscillations in the cerebral vasculature, with a spatial correlation length of ∼2 mm for arterioles. In complementary data from awake human subjects, the BOLD signal is spatially correlated among sulcus veins and specified intracortical veins of the visual cortex at similar ultra-slow rhythms. These data support the use of fMRI to resolve functional connectivity at the level of single vessels.

Keywords: BOLD; Calcium; GCaMP; cerebral blood volume; fMRI; functional connectivity; oscillation; resting state; single vessel; vasomotion.

Figure 1. Balanced Steady-State Free Precession (bSSFP)-based task-related single-vessel BOLD/CBV fMRI

(A) An A-V map shows individual venules (dark dots, blue markers) and arterioles (bright dots, red markers) in a 2D slice. (B) The BOLD fMRI map (left panel) and the semi-transparent map overlaid on the A-V map demonstrates the venule-dominated peak BOLD signal with the on/off block time series from a single venule ROI. (C) The CBV fMRI map (left panel) and the semi-transparent map overlaid on the A-V map show the arteriole-dominated peak CBV signal with the on/off block time series from a single arteriole ROI. (D) The averaged BOLD (left)/CBV(right) fMRI response function from venule (blue) and arteriole (red) voxels (n = 5 rats, mean ± s.e.m).

Figure 2. Using bSSFP-based rs-fMRI to map vascular-specific correlation patterns

(A) The A-V map shows individual penetrating arterioles and venules (blue arrowheads, venules; red arrowheads, arterioles). (B) The seed-based BOLD rs-fMRI correlation maps (0.01 – 0.1 Hz; seeds: cyan crosshairs) of two venule seeds (V1 and V2; left panel) and CBV rs-fMRI correlation maps (0.01 – 0.1 Hz; seeds: cyan crosshairs) of two arteriole seeds (A1 and A2; right panel). The lower panel is the BOLD signal time course of the two venule seed ROIs and two arteriole seed ROIs. (C) The seed-based CBV rs-fMRI correlation maps (0.01 – 0.1 Hz; seeds: cyan crosshairs) of two venule seeds (V1 and V2; left panel) and CBV rs-fMRI correlation maps (0.01 – 0.1 Hz; seeds: cyan crosshairs) of two arteriole seeds (A1 and A2; right panel). The lower panel is the CBV signal time course of the two venule seed ROIs and two arteriole seed ROIs. (D) The PSD of the venule and arteriole-specific resting-state BOLD (upper panel) and CBV (lower panel) fMRI time courses.

Figure 3. Vascular dynamic network connectivity in rats (14.1T)

(A) The A-V map of one representative rat (arteriole ROIs in red and venule ROIs in blue). (B–C) Scatter plots of the correlation coefficient (CC) of BOLD (B) and CBV (C) fMRI from venule-to-venule (V-V) pairs, arteriole-to-arteriole (A-A) pairs as the function of the inter-vessel distance from one representative rat. (D–E) The correlation matrices of all vessel pairs for the BOLD (D) and CBV (E) fMRI from one representative rat. (F–G) The mean CC value of the BOLD signal from the venule pairs is significantly higher than that of the arteriole pairs with large spatial inter-vessel distance (> 5 mm) (F, n = 5 rats, mean ± s.e.m, *, paired t-test, p < 0.03). In contrast, the mean CC value of the CBV signal from the arteriole pairs is significantly higher than that of the venule pairs with small spatial inter-vessel distance (~ 2 mm). (G, n = 5 rats, mean ± s.e.m, *, paired t-test, p < 0.03). (H–I) The averaged coherence graph of paired venules and arterioles from BOLD/CBV fMRI (H, BOLD fMRI, n = 5 rats, I, CBV fMRI, n = 5 rats, mean ± s.e.m). (J) The mean BOLD coherence coefficient of the venule pairs is significantly higher than that of arteriole pairs at the low-frequency range (0.01–0.04 Hz). (n=5 rats, paired t-test, **, p = 0.0009). (K) The mean CBV coherence value of paired venules is significantly lower than that of paired arterioles at the low-frequency range (0.01–0.04 Hz) (n = 5 rats, paired t-test, **, p = 0.007).

Figure 4. Correlation analysis of the single-vessel BOLD/CBV fMRI with GCaMP6f-mediated calcium signal

(A) The coronal view of the anatomical MR image with the optic fiber targeting the vibrissa S1 (upper panel). The A-V map from a 2D slice covering the deep cortical layer (lower panel). (B) The seed-based BOLD correlation maps from one representative venule voxels (V1) overlaid on the A-V map. (C) The correlation map between the BOLD fMRI signal and the calcium signal (band-pass filter: 0.01 – 0.1 Hz). Inset is a representative color-coded lag time map between the calcium signal with the BOLD fMRI of individual venules (CC > 0.25). (D) The time courses of the BOLD fMRI signal from vessel voxels (V1: blue, solid line; V2: blue, dotted line) and the slow oscillation calcium signal (green). (E) The cross-correlation function of the calcium signal and BOLD fMRI signal of two representative venules (Ca-V1 and Ca-V2) with positive peak coefficients at the lag time 2–3 s. (F) The mean correlation coefficient of the calcium signal with the BOLD fMRI signal of venules is significantly higher than that of arterioles (n = 7 rats, mean ± s.e.m, paired t-test, ***, p = 2.5×10⁻⁵). (G) The histogram of venules with lag times varied from 0.5 to 6s (CC > 0.25) and mean lag time at 2.30±0.19 s. (n = 7 rats, mean ± s.e.m). (H) The seed-based correlation maps of CBV fMRI from one arterioles voxel (A1) overlaid on the A-V map (left). (I) The correlation map between the CBV fMRI and calcium signal (band-pass filter: 0.01 – 0.1 Hz), Inset is a representative color-coded lag time map between the calcium signal and the CBV fMRI signal of individual arterioles (CC < −0.25). (J) The time course of the CBV fMRI signal from arteriole voxels (red, solid and dotted lines) and the slow oscillation calcium signal (green). (K) The cross-correlation function of the calcium signal and CBV fMRI signal of two representative arterioles (Ca-A1 and Ca-A2) with negative peak coefficients at the lag time 1–2 s. (L) The mean correlation coefficient of the calcium signal with the CBV fMRI signal of arterioles is significantly higher than that of venules (n = 4 rats, mean ± s.e.m, paired t-test, ***, p = 0.0002). (M) The histogram of arterioles with lag times varied from 0.5 to 5 s (CC < −0.25). The mean lag time is 1.76 ± 0.14 s (n = 4 rats, mean ± s.e.m), which is significantly shorter than the lag times of the calcium and venule BOLD signal (BOLD, n = 7, CBV, n = 4, p = 0.025). (N) The schematic drawing of the spatial and temporal correlation patterns of the slow oscillation signal coupling from neurons to vessels.

Figure 5. The task-related and resting state single vessel fMRI mapping in awake human subjects (3T)

(A) A sagittal view of the human brain with a 2D EPI slice located in the occipital lobe. (B) An averaged EPI image shows the pial veins in sulci as dark dots. (C) The checkerboard visual stimulation-evoked BOLD functional map with peak BOLD signals located at pial veins. (D) The seed-based BOLD correlation maps (0.01–0.1Hz; seeds: two veins (V1 and V2)) demonstrate vessel-dominated patterns. (E) The magnified view of the averaged EPI image from one representative subject (vein ROIs, left hemisphere, blue, right hemisphere, cyan). (F–G) The time courses of two veins in the task related (F) and resting state (G) (0.01 – 0.1 Hz) conditions. (H) The coherence graph of paired veins exhibits coherent oscillation at the frequency range of 0.01 – 0.1 Hz significantly higher than the higher frequency range (0.1 – 0.2 Hz; n = 6 subjects, mean± s.e.m, **, paired t-test, p = 0.008). (I) The scatter plot of the correlation coefficient (CC) from intra-and inter-hemispheric vein pairs. (J) The mean CC of inter-hemispheric vein pairs with the inter-vessel distance between 5 – 8 cm is significantly higher than that of intra-hemispheric vein pairs with distance between 3 – 3.5 cm.(***, n = 6 subjects, mean± s.e.m, t-test, p = 0.0002) (K–L) The evoked functional (K) and resting-state correlation (L) maps were smoothed from 1 mm to 5 mm (FWHM).

Figure 6. The intracortical vascular dynamic mapping with 9.4 T

(A) The A-V map is acquired from a 2D slice across the occipital lobe. (B–C) The intra-cortical veins (arrows) in the magnified view of region 1 and region 2 in the A-V map (left panel). The right panel shows the correlation map based on the selected seeds (the intra-cortical veins: blue arrows) with highly correlated voxels detected on the other intracortical veins (white arrows) in the gray matter. (D–E) The seed-based correlation maps with Vein 1 (V1), Artery 1 (A1) as seeds, respectively (seeds: cyan crosshairs). (F) The coherence graph of paired veins (blue) and arteries (red) identified by the A-V map demonstrates the slow fluctuations from 0.01 to 0.1 Hz. (G) The mean coherence coefficients of the paired veins are significantly higher than that of the paired arteries at low frequency (0.01 – 0.1 Hz)(n = 6 subjects, mean± s.e.m, paired t-test, **, p = 0.0009).