Forebrain Deletion of the Vesicular Acetylcholine Transporter Results in Deficits in Executive Function, Metabolic, and RNA Splicing Abnormalities in the Prefrontal Cortex

Abstract

One of the key brain regions in cognitive processing and executive function is the prefrontal cortex (PFC), which receives cholinergic input from basal forebrain cholinergic neurons. We evaluated the contribution of synaptically released acetylcholine (ACh) to executive function by genetically targeting the vesicular acetylcholine transporter (VAChT) in the mouse forebrain. Executive function was assessed using a pairwise visual discrimination paradigm and the 5-choice serial reaction time task (5-CSRT). In the pairwise test, VAChTdeficient mice were able to learn, but were impaired in reversal learning, suggesting that these mice present cognitive inflexibility. Interestingly, VAChT-targeted mice took longer to reach criteria in the 5-CSRT. Although their performance was indistinguishable from that of control mice during low attentional demand, increased attentional demand revealed striking deficits in VAChT-deleted mice. Galantamine, a cholinesterase inhibitor used in Alzheimer's disease, significantly improved the performance of control mice, but not of VAChT-deficient mice on the 5-CSRT. In vivo magnetic resonance spectroscopy showed altered levels of two neurochemical markers of neuronal function, taurine and lactate, suggesting altered PFC metabolism in VAChT-deficient mice. The PFC of these mice displayed a drastic reduction in the splicing factor heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNPA2/B1), whose cholinergic-mediated reduction was previously demonstrated in Alzheimer's disease. Consequently, several key hnRNPA2/B1 target transcripts involved in neuronal function present changes in alternative splicing in VAChT-deficient mice, including pyruvate kinase M, a key enzyme involved in lactate metabolism. We propose that VAChT-targeted mice can be used to model and to dissect the neurochemical basis of executive abnormalities.

Cholinergic Regulation of hnRNPA2/B1 Translation by M1 Muscarinic Receptors

“…Interestingly, we observed no change in hnRNPA2/B1 protein levels in striatum-specific VAChT mutants, whereas there is a body of evidence that cholinergic signaling can affect the levels of this protein in both cortical and hippocampal regions in vivo (Berson et al, 2012;Kolisnyk et al, 2013a). Given these findings, it is likely that it is an intrinsic property of the target cells themselves that dictate their change in hnRNPA2/B1 translation in response to cholinergic activity.…”

“…ChAT-ChR2-EGFP mice overexpress VAChT in the hippocampus and consequently present increased cholinergic tone and ACh release (Kolisnyk et al, 2013b). These mice present increased hnRNPA2/B1 protein levels (Fig.…”

“…Cholinergic tone can modulate signal processing by changing electrical properties of cells and by modulating intracellular sig-naling (Dajas-Bailador and Wonnacott, 2004;Soreq, 2015). In addition, it has become clear that cholinergic signaling can also regulate long-term gene expression, miRNAs (Shaked et al, 2009;Soreq, 2015), and alternative splicing (Berson et al, 2012;Kolisnyk et al, 2013a), all of which can modulate functional properties of cells (Blencowe, 2006;Novarino et al, 2014).…”

“…In addition, it has become clear that cholinergic signaling can also regulate long-term gene expression, miRNAs (Shaked et al, 2009;Soreq, 2015), and alternative splicing (Berson et al, 2012;Kolisnyk et al, 2013a), all of which can modulate functional properties of cells (Blencowe, 2006;Novarino et al, 2014). A key protein that regulates splicing events in AD is the heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1, which is reduced in brains of AD patients due to cholinergic deficiency (Berson et al, 2012;Kolisnyk et al, 2013a) and is reduced in mice with a conditional deletion of the vesicular acetylcholine (ACh) transporter in the forebrain (VAChT) (Kolisnyk et al, 2013a).…”

The NEWMEDS rodent touchscreen test battery for cognition relevant to schizophrenia

“…When reducing vesicular acetylcholine transporter (VAChT) in the prefrontal cortex specifically, a more subtle profile emerged in which VAChT-deficient mice showed impaired acquisition, and increased omissions at short stimulus duration probes of high attentional demand (Kolisnyk et al 2013b). When VAChT was overexpressed, mice displayed a profile of both impaired attention and impulsivity at shorter stimulus durations, with reduced accuracy and increased premature responding, respectively (Kolisnyk et al 2013a, b). The task has also been used to assess a model of autism, the inbred BTBR T+tf/J mouse, which also showed reduced levels of prefrontal acetylcholine (McTighe et al 2013).…”