Spatial Light Modulation – Two Photon Microscopy (SLM-2PM) allows to acquire fluorescence signals simultaneously from multiple neurons in thick samples.
This custom made setup, developed in collaboration with the physics department of Milano-Bicocca University, employs a Spatial Light Modulator to acquire three dimensional two photon images with a completely holographic method, without any moving part. The system also allows for the simultaneous recording of millisecond-scale optical signals from multiple neuronal elements.
We use 2PM-SLM to study, on a single cell scale, the activity of cerebellar microcircuits, through the recording of fluorescence signal from acute cerebellar slices stained with Calcium sensitive dyes, such as Fura-2.
Example of the image generated by Fura-2AM bulk loading of a cerebellar slice:
Multi-spot illumination was generated by pointing the laser beamlets toward selected neurons. The Ca2+ signal variations detected from the soma of GrCs in response to mossy fiber stimulation (10 impulses at 50 Hz) appear as a DF/F0 reduction induced by intracellular calcium increase in GrCs. The traces on the right show the time-course of DF/F0 at variable sampling frequencies (up to 1kHz) taken from a representative GrC.