Fig. 2: Validation of DATCre-Tg2576 model, a new mouse model for optogenetic stimulation of VTA in 6-month-old Tg2576 mice.

A Heterozygous DATCre female mice were crossed with heterozygous Tg2576 males to obtain DATCre/WT and DATCre/Tg2576 (Tg) mice for optogenetic stimulation of VTA DA neurons. B 6E10 immunostaining of intracellular Aβ levels in the hippocampus of 6-month-old DATCre/WT and DATCre/Tg2576 mice. Nuclei are counterstained with DAPI and neurons with NeuroTrace (scale: 250 µm; inset 25 µm). C-D Representative coronal sections of the midbrain showing immunofluorescence for TH⁺ neurons in DATCre/WT and DATCre/Tg mice at 6 months of age (n = 4 mice / group; scale bar: 100 µm; inset 50 µm) and stereological TH⁺ cell count in total VTA (Unpaired t-test: **p = 0.0015) and unilateral SNpc (D; scale bar: 50 µm). E Confocal images and plot showing TH⁺ fibre density (expressed as fibres / 250 μm; scale bar: 50 µm) in the dorsal hippocampus of DATCre/WT and DATCre/Tg mice at 6 months of age (n = 4 mice / group. Unpaired t-test: **p = 0.0080). Nuclei are counterstained with DAPI. F 5-month-old DATCre/WT mice were bilaterally infused in the midbrain with AAV-EF1a-DIO-hChR2(H134R)-eYFP for selective expression of the ChR2-eYFP fused protein in the mouse midbrain. Mice were used for experiments 1-month post-injection. G Representative confocal images show eYFP fluorescence in a brain slice used for electrophysiology. The square indicates the midbrain (scale bar: 500 µm). The images on the right show the co-labelling of eYFP and TH (scale bar: 10 µm). H Representative confocal images of eYFP fluorescence in the dorsal hippocampus of a 6-month-old DATCre/WT mouse injected with the viral construct in the midbrain (scale bar: 25 µm). Nuclei are counterstained with DAPI and neurons with NeuroTrace. I Schematic representation of optogenetic stimulation of dopaminergic (eYFP⁺) neurons in VTA of 6-month-old DATCre/WT applied during electrophysiology recordings. Representative traces of spontaneous firing of a DA neuron in the lateral VTA at 6 months of age before photostimulation, and during phasic (shown in detail in the inset) and prolonged light stimulation (scale bar: 100 ms). Note how the spontaneous pacemaking activity is restored following the end of phasic stimulation, but the neuron remains silent during the prolonged stimulation. [Schematics were created using BioRender (https://biorender.com)].