optogenetics experiments require illumination or light stimulation of light sensitive neurons in specific areas of the brain. The light is introduced in brain via fiber-optic cannulas which consist of receptacle and optical fiber whose tip protrudes inside the brain. When the optical fiber is implanted in brain its tip delivers narrow cone of light in front of the fiber. The angular spread of light coming from a fiber tip is determined by the numerical aperture (NA) of the fiber. For example, the optical fiber with 0.22 NA spreads the light within the apex angle of 25.4° in air . When implanted or immersed into body liquids the angular spread of the light coming out of the fiber tip is reduced by the factor of 1.33 to 19.1°, where 1.33 is approximately the refractive index of the body liquids. As this angle is relatively small, some of the neurons of interest for optogenetics or electrophysiology experiments may not be illuminated.
To overcome this limitation, we have added diffuser layer on the tip of the optical fiber to increase its angular light spread. Typically, our fiber-optic cannulas with diffuser layer fill the 300° apex angle (solid angle > 3.7π [sr]) with light. This is over 74 times larger in terms of solid angle, when compared with 0.22 NA standard fiber-optic cannula where the solid angle is 0.05π [sr]). In air, the light intensity falls-off gradually as the angle increases (Figure 1). Consequently, for the same amount of input light power, the diffusive cannula will illuminate larger area albeit with reduced illumination intensity. To estimate the illumination pattern in biological tissues, measurement was also done in water. Water modify the light transmission at tip and yield to a significant larger output intensity than in air; total output power is 1.56 ± 0.05 X bigger in water (mean ± S.D., N=7 tested cannulas). The resulting illumination pattern is maximum in front of the tip and intensity never drop under 20% of peak maximum over 300° (Figure 1 and 2). The intensity drops theoretically as 1/r² where r is the distance from fiber tip. Additionally, the loss induced by the diffuser are almost 50% of the input power in air yet drop around to 25% in water. Therefore, the diffusive tip enlarges the illumination area but needs higher illumination power.
Figure 1: A)
Photo of the optical fiber tip with diffuser layer. B)
Experimental angular output light intensity from a flat ended optical fiber (0.22 NA, 200µm core diameter) and a similar one with a diffuser. Result are given for the same tips in air (black) and in water (green). C-D)
Experimental angular distribution of the output light intensity for the fibers used in B
-flat ended optical fiber, D
-optical fiber with diffuser). Arrow represents the output intensity and shows that the front output intensity is around 70 times smaller with the diffuser compare to flat ended fiber.
Illumination patterns of two fiber-optic cannulas with 200 µm 0.22 NA optical fiber and blue light (λ=470nm). The tips of both fibers are polished at right angle, while the tip of the lower fiber has diffuser overlay. A)
Two optical fiber tips immersed in a fluorescent solution (water diluted fluorescein, FITC). The excited fluorescence is observed as green light. B)
Two optical fiber tips in air with smoke screen to render the beam visible. C)
Microscope view of both illuminations when fibers were laid on a white screen.
Advantages of fiber-optic cannula with diffuser layer:
- light is spread into larger solid angle, over 70 X bigger than conventional optical fiber,
- several electrodes can be illuminated,
- the illumination pattern allows very efficient cross-section excitation,
- the rapid intensity drop useful for local stimulation; varying input power enables fine tuning of excitation reach.
- the wide illumination requires two order of magnitude stronger input power to obtain the same light intensity,
- the inherent loss of the diffuser:50% in air and 25% in water.
- the diffusing layer is sensitive to certain solvent as acetone. Though, it can be cleaned with isopropanol alcohol.