Variable Neuronal Expression of Green Florescent Protein in the Brain of a Transgenic Swine Reporter Model

Transgenic pigs are emerging as large animal models for biomedical research. Green fluorescent protein (GFP)-expressing transgenic reporter pigs (GFP-Tg; NSRRC:0016 GFP NT92) express GFP via the ubiquitous pCAGG promoter. Their level of GFP expression across brain regions has not yet been evaluated. Brain tissue sections from GFP-Tg animals were immunostained against GFP and the neuronal marker NeuN, or against GFP, NeuN, and the dopaminergic neuronal marker tyrosine hydroxylase (TH). The presence and intensity of neuronal GFP expression was evaluated in the amygdala, caudate, hippocampus, primary motor cortex (M1), putamen, and substantia nigra pars compacta (SNpc). Images from each region were captured, and individual neurons were counted and rated based on their intensity of GFP expression as being GFP positive (GFP+), equal (GFP=) to neuropil, dim (GFPd), or negative (GFP-). Nearly all neurons in the amygdala (99.2%), caudate (99.0%), hippocampus (99.7%), M1 (99.4%), and putamen, (98.8%) showed high GFP-immunoreactivity (combined GFP+ and GFP=), in contrast to the SNpc, which had significantly less (61.4%; p=0.0266). The number of GFP- neurons was significantly higher in the SNpc than in the amygdala (p =0.0342), hippocampus (p=0.0366), and M1 (p=0.0437). Overall, this study demonstrates high neuronal GFP expression in the GFP-Tg reporter pig, but with variability between brain regions, particularly decreased GFP expression in dopaminergic nigral neurons. Future research utilizing this model, such as studies to evaluate the efficacy of genome-editors for Parkinson's disease through their ability to turn off GFP expression in the nigrostriatal system, should account for this neuronal GFP-expression variability.