Toward microendoscopy-inspired cardiac optogenetics in vivo: technical overview and perspective

Abstract

The ability to perform precise, spatially localized actuation and measurements of electrical activity in the heart is crucial in understanding cardiac electrophysiology and devising new therapeutic solutions for control of cardiac arrhythmias. Current cardiac imaging techniques (i.e. optical mapping) employ voltage- or calcium-sensitive fluorescent dyes to visualize the electrical signal propagation through cardiac syncytium in vitro or in situ with very high-spatiotemporal resolution. The extension of optogenetics into the cardiac field, where cardiac tissue is genetically altered to express light-sensitive ion channels allowing electrical activity to be elicited or suppressed in a precise cell-specific way, has opened the possibility for all-optical interrogation of cardiac electrophysiology. In vivo application of cardiac optogenetics faces multiple challenges and necessitates suitable optical systems employing fiber optics to actuate and sense electrical signals. In this technical perspective, we present a compendium of clinically relevant access routes to different parts of the cardiac electrical conduction system based on currently employed catheter imaging systems and determine the quantitative size constraints for endoscopic cardiac optogenetics. We discuss the relevant technical advancements in microendoscopy, cardiac imaging, and optogenetics and outline the strategies for combining them to create a portable, miniaturized fiber-based system for all-optical interrogation of cardiac electrophysiology in vivo.

Fig. 1

Example of a fiber-optic imaging system for multisite optical recording of cardiac tissue. The fiber bundle consists of multimode fibers terminated at different lengths in order to Di-4-ANBDQBS. Collected light from the fiber bundle is either imaged onto a second fiber bundle connected to individual photodiodes to perform ratiometric measurements^, or imaged directly on to a photodiode array (PDA).^,,,

Fig. 2

Schematic of early microendoscopic imaging systems. (a) Fiber-optic imaging bundle (FOIB) system: Light is scanned across a FOIB using a two-dimensional (2-D) scanner and focused on to the sample using an objective to perform the single-photon confocal imaging. (b) Compact two-photon imaging system: A probe containing a lens system scans light across a sample to perform the two-photon imaging in vivo and in vitro. ^,, (c) Miniaturized microscope: Light from a single-mode fiber is scanned across a sample using a head-mounted unit containing a scanner, dichroic mirror, focusing lenses, and photomultiplier tube (PMT) to perform in vivo two-photon imaging.

Fig. 3

Microendoscopic systems using gradient index (GRIN) lenses. (a) Scanning FOIB system: Excitation light is scanned across an FOIB and focused on the sample using a GRIN doublet to perform either single-photon confocal imaging or two-photon imaging in vitro. (b) Minimally invasive microendoscope using GRIN system: A focusing lens, GRIN doublet, and dichroic mirror can be incorporated in a head-mounted unit to perform in vivo single-photon imaging in rat brain^, or in vivo two-photon imaging.^,, (c) Lensed fiber endoscope: Light from a lens fiber is focused onto a sample using a GRIN lens system and scanned across the sample using a raster scanner to perform the single-photon imaging. (d) Light is delivered using a fiber or fiber bundle to a GRIN system which can be mounted on the head of freely moving mice to perform in vivo single-photon imaging. (e) Dual-axis confocal microscope: Light is collimated from a single-mode fiber using a GRIN coupler and is focused on to a MEMS system using a parabolic mirror. It is then scanned across a sample using a GRIN-based endoscopic probe to perform in vitro and in vivo single-photon confocal imaging.^, (f) Portable microendoscope system: A probe consisting of a GRIN doublet fused to a dichroic microprism that can be mounted on to the head of freely moving mice to perform in vivo two-photon imaging.^,

Fig. 4

Schematic of endoscopic paths to access the electrical conduction system of the heart. Blue paths depict the access via the right side of the heart, where a catheter is inserted through the femoral vein to access the right atrium via the inferior vena cava (IVC) or through the brachial vein via the superior vena cava (SVC) (a and b). From the right atrium, the catheter can then access the atrioventricular (AV) node, sinoatrial (SA) node, and bundle of His, or be guided into the coronary sinus (CS) or right ventricle. Pink paths depict the access via the left side of the heart, where a catheter is inserted through the femoral artery in order to access the aorta (c); from there the SA node can be accessed via the coronary arteries or the catheter can be guided into the left ventricle.

Fig. 5

Optical path diagram of proposed fiber-optic based microendoscopic device for cardiac optogenetics. Excitation light for the fluorescent dye is combined with stimulation light using a dichroic mirror, and both beams are coupled into an optical fiber using a fiber coupler. Light from the fiber is focused on to the sample using an endoscopic probe. The fluorescent signal is then coupled back into the fiber using the same probe and sent to the detector using a dichroic mirror.

Fig. 6

Scattering and absorption data for rat heart tissue. (a) Absorption and reduced scattering coefficients for visible wavelengths in the rat heart (adapted from Ref. 122); (b) effective attenuation coefficients for visible wavelengths in rat heart tissue (adapted from Ref. 122) and diluted blood at 100% ${\mathrm{O}}_2$ saturation (adapted from Ref. 123).