Unlocking the Power of Non-linear and Ultrafast Optics in Biology: Applications to New Tools for Bio-imaging and Neuroscience

Alipasha Vaziri
Howard Hughes Medical Institute

In the recent years from the intersection of physics and biology, significant advancements in life sciences have emerged. This development has been fueled by two main drivers. On one hand many biological fields such as neuroscience are currently limited by the available tools; hence the development of new physical techniques and methods have enabled new biological discoveries. On the other hand, a physics-based approach to addressing biological questions can lead to an understanding of biological problems on a more fundamental level. In this context I will discuss our recent application of non-linear and ultrafast optics to the development of new techniques in functional and structural bio-imaging and report on two advancements; one in optical super-resolution imaging and another in subcellular optical control of neuronal activity with high temporal precession.

A general feature of most of the super-resolution imaging techniques based on photo-activated localization microscopy (PALM) has been that the imaging depth is limited to a fraction of an optical wavelength. However, to study whole cells, the extension of these methods to a 3D-super-resolution technique is required. We have overcome this limitation by using a two-photon illumination technique called temporal focusing in which the spectral properties of the pulse are used to control its axial intensity distribution in space. Using temporally focused beams we have demonstrated super-resolution imaging in 3D over an axial range of ~10μm in various biological samples.

The combination of genetic tools and optics have for the first time provided a method for cell type specific initiation of neuronal response in vitro and in vivo with a wide range of neurobiological applications. However in almost all studies widefield light sources combined with linear excitation of molecules have been used leading to a spatially unlocalized activation of a large neuronal population. Moreover, excitation via two-photon scanning which usually provides higher optical localization has proven to be challenging for the excitation of Channelrhodopsin, the most widely used light gated ion-channel. Using two-photon “sculpted light” activation of Channelrhodopsin we have demonstrated targeted, single cell and subcellular specific optogenetic initiation of neuronal response. The unprecedented spatial and temporal resolution of this method has a wide range of applications in fundamental neuroscience questions that could have not been addressed until now by the current methods. The extension of light sculpting to more general spatial light distributions in 3D will have further number of important applications in structural and functional bio-imaging.