Author:

Abstract:

The response of mammalian visual cortex to enucleation or retinal lesioning is a well-documented model for cortical plasticity. Differential molecular distribution profiles are routinely explored by in situ hybridization or immunocytochemistry. We here introduce MALDI MSI as an exquisite, fully complementary method to current molecular investigations in neuroscience, bypassing the need for a reporter system (stains or labels) or prior knowledge of molecules involved in brain plasticity. Coronal 10 µm mouse visual cortex sections were transferred to a stainless steel MALDI target, freeze-dried, washed (5s 70% Eth, 95% Eth) and sinapinic acid matrix deposited using aerosol spray with a coaxial N2 gas flow (flow-rate at 30 µl/min). Spectra from 4-25 kDa were acquired on a Bruker Reflex IV MALDI-TOF device operated in linear mode. Laser energy attenuation was at 50%. Sixty shots were accumulated per pixel position. Two software tools had to be developed for the Bruker Reflex IV: the first creates a custom raster pattern to the specifications of the user, according to the dimensions and position of the section and allows for automated pixel-by-pixel acquisition of spectra using the AutoXecute routines. The second reads the resulting spectra, converts them to a file in the Analyze 7.5 image format that can be imported in BioMap software for image interpretation. Upon method optimization (reproducibility, resolution), brain plasticity-induced protein profiles were examined by comparing retinally lesioned, enucleated and normal mouse brain MS images. For internal validation of distribution profiles, both hemispheres were imaged. Adjacent Nissl-stained 25 µm sections were compared with or overlaid on the MALDI MS images in BioMap to provide an anatomical reference view. We effectively introduced the high-quality application of MALDI MSI in neuroscience as an innovative research strategy in ongoing molecular investigations of brain plasticity.