Laser probes rat's brain in vivo

17 Jun 2002

German and US researchers image the brain of a moving rat with a miniature two-photon microscope.

Neurotechnique

Researchers from the Max-Planck Institute for Medical Research in Germany and Bell Labs in the US have developed a miniaturized microscope based on two-photon fluorescence excitation. Winfried Denk and colleagues use this device for imaging the brain of a moving rat (Neuron 31 903).

Blood-carrying capillaries and dendrite activity were imaged by labelling compounds with a fluorescent agent that was injected into the bloodstream. Dendrites are the tree-like structures in which the cell collects the inputs from other neurons. It is also believed that this is where memory is stored.

Imaging in motion

One of the advantages of using two-photon fluorescence is the depth of penetration into the tissue that can be achieved. Using this technique, the researchers have acquired images of neural structures buried up to 250µm into the brain. "Many interesting neurobiological phenomena occur in the first 200µm," said Denk.

The 25g microscope was attached to the head of the rat and a cranial window was used to image the brain. Images of the fluorescence were recorded using a commercially available compact photomultipler tube, owing to the small amount of fluorescence emitted.

Two-photon fluorescence is only generated at the beam waist of the focused laser. In this instance, the beam waist diameter was approximately 1µm and matched the size of a typical dendrite, allowing imaging to be performed at a rate of two images per second. The researchers performed larger area scans and implemented an efficient mechanism for scanning the beam waist.

Denk believes his group is the first to develop a microscope capable of monitoring brain activity in an animal that is free to move and behave in its natural environment. The group plans to reduce the size of the microscope even further and concentrate on increasing the excitation efficiency of the fluorescence. "The ultimate goal is functional imaging of neurons in freely moving animals," said Denk.