PMC

(A) Schematic of co-registered optical coherence tomography (OCT) and optical intrinsic signal imaging (OISI) system. TIA: Transimpedance amplifier (TIA). (B) Photograph showing the OCT sample arm combined with CCD field of view through the dichroic hot mirror.

Representative responses from ROIs on five different animals. Each color represents the results from one animal. Both positive (solid line) and negative (dashed line) responses are presented. Although temporal location and intensity of the peak varied slightly from animal to animal, in general, the response corresponded well with the stimulation pattern.

Spatial-temporal evolution of functional OCT signals in the rat cortex. A fractional change map demonstrates the presence of positive (warm colors) and negative (cool colors) changes in OCT signals during stimulation. Functional OCT images at each individual temporal point (t = 1–15 s) reveal the presence of highly localized regions of activation in the cortex that persist throughout stimulation. Bar: 500 μm.

(A) Localization of activation area in OISI. Two OCT scans are denoted by arrows “1” and “2”. (B) Functional OCT image at time window 4–6 s over the activation region denoted by arrow “1” during the stimulation of the contra-lateral forepaw. (C) Functional OCT image at time window 4–6 s over the scan denoted by arrow “2” during the stimulation of the contra-lateral forepaw. The activation signal is significantly less than in region “1”, which agrees with the OISI signals in (A). (D) Functional OCT image at time window 4–6 s over the scan denoted by arrow “1” during the stimulation of the ipsi-lateral forepaw. Note that no activation signal is detected.

Axial and transverse line profiles of functional OCT signals. (A) Position of the axial lines (x1 and x2) and transverse lines (z1 and z2) on OCT structural (left) and functional (right) images. (B) Left: Plot of differential functional signal along line x1. Right: Plot of raw linear signals at both baseline (pre-stimulation) and during stimulation period along line x1. Inset shows enlarged (×1.5) view of region denoted by “1”. Black arrow indicates the edge of skull. (C) Left: Plot of differential functional signal along line x2. Right: Plot of raw linear signals at both baseline (pre-stimulation) and during stimulation period along line x2. Inset shows enlarged (×2) view of region denoted by “2”. Black arrow indicates the edge of skull. (D) Left: Plot of differential functional signal along line z1. Right: Plot of raw linear signals at both baseline (pre-stimulation) and during stimulation period along line z1. Black arrow indicates the edge of skull. (E) Left: Plot of differential functional signal along line z2. Right: Plot of raw linear signals at both baseline (pre-stimulation) and during stimulation period along line z2.

Swept-source OCT image of functional activation. (A) Corresponding OISI image showing activation area (denoted by blue color); (B) En face OCT image; (C) Cross-sectional OCT imaging showing regions of functional activations. OCT image in (C) is acquired along the plane indicated by the red lines in both (A) and (B) (For interpretation of the references to color in this figure legend, the reader is referred to the web version of the article).

(A) Structural OCT image with two regions indicated by red and blue boxes. (B) The time-courses of the functional OCT for these two regions. (C) Spatially-resolved time-lags overlaid with structural OCT image. Different colors indicate different time-lags with respect to the averaged OISI time-course. (D) The histogram of the time-lags. (E) Temporal plots of averaged signal responses with time lags at different windows (For interpretation of the references to color in this figure legend, the reader is referred to the web version of the article).

(A) The functional OCT image (red-blue color scale) at time window 4–6 s with significance level α < 0.001 superimposed with the OCT anatomical image (grey scale); (B) plots of positive, negative, and summation OCT signals at regions with significance level α < 0.001. Note that a net positive change occurs during the activation period. The time course of OISI signal (×10) is also included to show the temporal correspondence (For interpretation of the references to color in this figure legend, the reader is referred to the web version of the article).

(A) Structural OCT image overlaid with the time-lags with respect to the averaged OISI time-course at different activation regions. The time-lags are color-coded (unit: second). (B) Plot of the averaged time-lags at the same depth (from the skull surface) versus the depth. Two distinct regions are indicated by different slopes (S1 and S2). (C) Box plot of the S1 and S2 from 9 independent runs on 5 rats showing statistically significant difference (*p < 0.0001). (D) Histology of rat somatosensory cortex indicating layered structures (layer I, II, III are specified).

Precise co-registration of OCT imaging to the region of functional activation. (A) The OCT scan is directed across the region of interest (indicated by blue double-head-arrow) as measured with OISI. Anatomical features such as blood vessels (V) can be visualized in OISI and can be used to co-register with OCT image. (B) Structural OCT image reveals high-resolution visualization of the skull (S), surface vasculature (V), and meningeal layers, including the dura mater (D) and arachnoids (A). Cortex region (C) can also be visualized. (C) Three-dimensional rendering of the volume acquired by OCT. (D) En face OCT image reveals the characteristic blood vessel network which can be used to register the OCT imaging volume with OISI.

Effects of different averaging window sizes. (A) 1 × 1; (B) 3 × 3; (C) 7 × 7. The histograms of the signals resulting from these three averaging windows are shown in (D). Bar: 500 μm. (E) Non-averaged OCT raw signal (normalized to baseline) trace showing functional responses (blue curve) corresponding to the stimulation trial (beginning of each trial is indicated by the red curve). (F) The Fourier transform of the signal trace revealing the stimulation frequency (0.05 Hz, indicated by the red arrow) and its harmonics. The inset shows an enlarged view (For interpretation of the references to color in this figure legend, the reader is referred to the web version of the article).

Representative results of OISI and OCT imaging of functional activation. (A) En face functional activation map from OISI with co-registered OCT scan indicated by the red arrow. (B) The averaged time-course for OISI over the region denoted by dashed box in (A) with stimulation period shaded grey. (C) Functional OCT image superimposed with the structural image. In order to simultaneously display these two images, we threshold the functional OCT signals to those with responses > 5% (or <−0.5%). (D) Representative functional OCT time-courses for regions of interest specified by solid arrow (positive changes) and hollow arrow (negative changes). (E) Map of p values showing the statistical significance of the fractional OCT changes from the baseline. (F) The lateral line profile of OISI signal changes and the depth-integrated OCT signal changes showing the spatial correspondence of OCT and OISI signals. The OISI and OCT signal changes are scaled to overlap in order to provide better comparison.