Extracting surface activation time from the optically recorded action potential in three-dimensional myocardium.

Optical mapping has become an indispensible tool for studying cardiac electrical activity. However, due to the three-dimensional nature of the optical signal, the optical upstroke is significantly longer than the electrical upstroke. This raises the issue of how to accurately determine the activation time on the epicardial surface. The purpose of this study was to establish a link between the optical upstroke and exact surface activation time using computer simulations, with subsequent validation by a combination of microelectrode recordings and optical mapping experiments. To simulate wave propagation and associated optical signals, we used a hybrid electro-optical model. We found that the time of the surface electrical activation (t(E)) within the accuracy of our simulations coincided with the maximal slope of the optical upstroke (t(F)*) for a broad range of optical attenuation lengths. This was not the case when the activation time was determined at 50% amplitude (t(F50)) of the optical upstroke. The validation experiments were conducted in isolated Langendorff-perfused rat hearts and coronary-perfused pig left ventricles stained with either di-4-ANEPPS or the near-infrared dye di-4-ANBDQBS. We found that t(F)* was a more accurate measure of t(E) than was t(F50) in all experimental settings tested (P = 0.0002). Using t(F)* instead of t(F50) produced the most significant improvement in measurements of the conduction anisotropy and the transmural conduction time in pig ventricles.

Probing field-induced tissue polarization using transillumination fluorescent imaging.

Despite major successes of biophysical theories in predicting the effects of electrical shocks within the heart, recent optical mapping studies have revealed two major discrepancies between theory and experiment: 1), the presence of negative bulk polarization recorded during strong shocks; and 2), the unexpectedly small surface polarization under shock electrodes. There is little consensus as to whether these differences result from deficiencies of experimental techniques, artifacts of tissue damage, or deficiencies of existing theories. Here, we take advantage of recently developed near-infrared voltage-sensitive dyes and transillumination optical imaging to perform, for the first time that we know of, noninvasive probing of field effects deep inside the intact ventricular wall. This technique removes some of the limitations encountered in previous experimental studies. We explicitly demonstrate that deep inside intact myocardial tissue preparations, strong electrical shocks do produce considerable negative bulk polarization previously inferred from surface recordings. We also demonstrate that near-threshold diastolic field stimulation produces activation of deep myocardial layers 2-6 mm away from the cathodal surface, contrary to theory. Using bidomain simulations we explore factors that may improve the agreement between theory and experiment. We show that the inclusion of negative asymmetric current can qualitatively explain negative bulk polarization in a discontinuous bidomain model.

Cerebral microbleeds in adult survivors of childhood acute lymphoblastic leukemia treated with cranial radiation.

Cranial radiation therapy is associated with white matter-specific brain injury, cortical volume loss, mineralization, microangiopathy and neurocognitive impairment in survivors of childhood acute lymphoblastic leukemia. In this retrospective cross-sectional analysis, neurocognitive testing and 3 T brain MRI's were obtained in 101 survivors treated with cranial radiation. Small focal intracerebral hemorrhages only visible on exquisitely sensitive MRI sequences were identified and localized using susceptibility weighted imaging. Modified Poisson regression was used to assess the effect of cranial radiation on cumulative number and location of microbleeds in each brain region, and multiple linear regression was used to evaluate microbleeds on neurocognitive outcomes, adjusting for age at diagnosis and sex. At least one microbleed was present in 85% of survivors, occurring more frequently in frontal lobes. Radiation dose of 24 Gy conveyed a 5-fold greater risk (95% CI 2.57-10.32) of having multiple microbleeds compared to a dose of 18 Gy. No significant difference was found in neurocognitive scores with either the absence or presence of microbleeds or their location. Greater prevalence of microbleeds in our study compared to prior reports is likely related to longer time since treatment, better sensitivity of SWI for detection of microbleeds and the use of a 3 T MRI platform.

Spontaneous onset of atrial fibrillation.

Most commonly, atrial fibrillation is triggered by rapid bursts of electrical impulses originating in the myocardial sleeves of pulmonary veins (PVs). However, the nature of such bursts remains poorly understood. Here, we propose a mechanism of bursting consistent with the extensive empirical information about the electrophysiology of the PVs. The mechanism is essentially non-local and involves the spontaneous initiation of non-sustained spiral waves in the distal end of the muscle sleeves of the PVs. It reproduces the experimentally observed dynamics of the bursts, including their frequency, their intermittent character, and the unusual shape of the electrical signals in the pulmonary veins that are reminiscent of so-called early afterdepolarizations (EADs).

Near-infrared voltage-sensitive fluorescent dyes optimized for optical mapping in blood-perfused myocardium.

BACKGROUND: Styryl voltage-sensitive dyes (e.g., di-4-ANEPPS) have been used successfully for optical mapping in cardiac cells and tissues. However, their utility for probing electrical activity deep inside the myocardial wall and in blood-perfused myocardium has been limited because of light scattering and high absorption by endogenous chromophores and hemoglobin at blue-green excitation wavelengths. OBJECTIVE: The purpose of this study was to characterize two new styryl dyes--di-4-ANBDQPQ (JPW-6003) and di-4-ANBDQBS (JPW-6033)--optimized for blood-perfused tissue and intramural optical mapping. METHODS: Voltage-dependent spectra were recorded in a model lipid bilayer. Optical mapping experiments were conducted in four species (mouse, rat, guinea pig, and pig). Hearts were Langendorff perfused using Tyrode's solution and blood (pig). Dyes were loaded via bolus injection into perfusate. Transillumination experiments were conducted in isolated coronary-perfused pig right ventricular wall preparations. RESULTS: The optimal excitation wavelength in cardiac tissues (650 nm) was >70 nm beyond the absorption maximum of hemoglobin. Voltage sensitivity of both dyes was approximately 10% to 20%. Signal decay half-life due to dye internalization was 80 to 210 minutes, which is 5 to 7 times slower than for di-4-ANEPPS. In transillumination mode, DeltaF/F was as high as 20%. In blood-perfused tissues, DeltaF/F reached 5.5% (1.8 times higher than for di-4-ANEPPS). CONCLUSION: We have synthesized and characterized two new near-infrared dyes with excitation/emission wavelengths shifted >100 nm to the red. They provide both high voltage sensitivity and 5 to 7 times slower internalization rate compared to conventional dyes. The dyes are optimized for deeper tissue probing and optical mapping of blood-perfused tissue, but they also can be used for conventional applications.

Paired self-compensated spin-lock preparation for improved T1ρ quantification.

PURPOSE: Spin-lock (SL) imaging allows quantification of the spin-lattice relaxation time in the rotating frame (T1ρ). B0 and B1 inhomogeneities impact T1ρ quantification because the preparatory block in SL imaging is sensitive to the field heterogeneities. Here, a modified preparatory block (PSC-SL) is proposed that attempts to alleviate SL sensitivity to field inhomogeneities in scenarios where existing approaches fail, i.e. high SL frequencies. METHODS: Computer simulations, phantom and in vivo experiments were used to determine the effect of field inhomogeneities on T1ρ quantification. Existing SL preparations were compared with PSC-SL in different conditions to assess the advantages and disadvantages of each method. RESULTS: Phantom experiments and computer modeling demonstrate that PSC-SL provides superior T1ρ quantification at high SL frequencies in situations where the existing SL preparation methods fail. This result has been confirmed in pre-clinical neuro and body imaging at 7T. CONCLUSION: PSC-SL complements existing methods by increasing the accuracy of T1ρ quantification at high spin-lock frequencies when large field inhomogeneities are present. A-priory information about the experimental conditions such, as field distribution and spinlock frequency are useful for selecting an appropriate spin-lock preparation for specific applications.

Imaging electrical excitation inside the myocardial wall.

Cardiac arrhythmias are often triggered by ectopic membrane depolarization originating deep inside the myocardial wall. Here we propose a new method utilizing a novel near-infrared voltage-sensitive fluorescent dye DI-4-ANBDQBS to determine the three-dimensional (3D) coordinates of the sources of such depolarization. We tested the method in live preparations of pig left and right ventricular myocardium (thickness 8-18 mm) and phantoms imitating the optical properties of myocardial tissue. The method utilizes an alternating transillumination approach that involves comparing pairs of simultaneously recorded broad-field epifluorescence and transillumination images produced at two alternating directions of illumination. Recordings were taken simultaneously by two CCD cameras facing the endocardial and epicardial surfaces of the heart at a frame rate up to 3 KHz. In live preparations, we were able to localize the origin of the depolarization wave with a precision of ±1.3mm in the transmural direction and 3 mm in the image plane. The accuracy of detection was independent of the depth of the source inside ventricular wall.

Monitoring intramyocardial reentry using alternating transillumination.

Intramyocardial reentry is implicated as a primary cause of the most deadly cardiac arrhythmias known as polymorphic ventricular tachycardia and ventricular fibrillation. However, the mechanisms involved in the triggering of such reentry and controlling its subsequent dynamics remain poorly understood. One of the major obstacles has been a lack of adequate tools that would enable 3D imaging of electrical excitation and reentry inside thick ventricular wall. Here, we present a new experimental technique, termed alternating transillumination (AT), aimed at filling this gap. The AT technique utilizes a recently synthesized near-infrared fluorescent voltage-sensitive dye, DI-4-ANBDQBS. We apply AT to study the dynamics of reentry during shock-induced polymorphic ventricular tachycardia in pig myocardium.

A novel near-infrared voltage-sensitive dye reveals the action potential wavefront orientation at increased depths of cardiac tissue.

Recently, novel near-infrared (NIR) voltage-sensitive dyes were developed for imaging electrical activity in blood-perfused hearts and for tomographic applications. However, their usefulness for conventional surface mapping is unclear. The spectral shift to the NIR range significantly increases the penetration depth of light into the tissue, thus increasing the intramural volume contributing to the optical action potential (OAP). Here, we characterize both computationally and experimentally the effect of increased penetration depth on the OAP upstroke, the OAP component most sensitive to optical scattering and absorption, and the activation maps. Optical imaging of cardiac electrical activity was performed in isolated rat hearts (n = 5) paced from the LV mid free wall. We used the NIR dye JPW-6033 (excitation at 660nm, acquisition at >695nm). The conventional dye DI-4-ANEPPS (excitation at 532nm, acquisition at 700 DF50nm) was used for comparison. To simulate OAP we utilized a hybrid model that couples light transport equations with the model of electrical propagation. As expected, the switch from DI-4-ANEPPS to JPW-6033 significantly increased the upstroke duration: from 3.95 + or - 0.69ms to 5.39 + or - 0.82 ms, respectively. However, activation maps were largely unaffected. The correlation between the shape of the optical upstroke, and the averaged subsurface wave front orientation was also preserved. The computer simulations are in excellent agreement with the experimental data. In conclusion, our analysis suggests that despite significant increase in upstroke duration, the novel NIR dyes can be a useful alternative to conventional dyes in surface mapping applications.

New near-infrared optical probes of cardiac electrical activity.

Styryl voltage-sensitive dyes (e.g., di-4-ANEPPS) have been widely and successfully used as probes for mapping membrane potential changes in cardiac cells and tissues. However, their utility has been somewhat limited because their excitation wavelengths have been restricted to the 450- to 550-nm range. Longer excitation/emission wavelength probes can minimize interference from endogenous chromophores and, because of decreased light scattering and lower absorption by endogenous chromophores, improve recording from deeper tissue layers. In this article, we report efforts to develop new potentiometric styryl dyes that have excitation wavelengths ranging above 700 nm and emission spectra extending to 900 nm. Three dyes for cardiac optical mapping were investigated in depth from several hundred dyes containing 47 variants of the styryl chromophores. Absorbance and emission spectra in ethanol and multilamellar vesicles, as well as voltage-dependent spectral changes in a model lipid bilayer, have been recorded for these dyes. Optical action potentials were recorded in typical cardiac tissues (rat, guinea pig, pig) and compared with those of di-4-ANEPPS. The voltage sensitivities of the fluorescence of these new potentiometric indicators are as good as those of the widely used ANEP series of probes. In addition, because of molecular engineering of the chromophore, the new dyes provide a wide range of dye loading and washout time constants. These dyes will enable a series of new experiments requiring the optical probing of thick and/or blood-perfused cardiac tissues.