Castaneda-Hernandez, G. | Search bioRxiv

bioRxiv2025

Adeno-Associated Virus-Induced Neurotoxicity is Prevented by CpG Depletion

Adeno-associated viruses (AAVs) are the vector of choice for gene delivery to the nervous system. While AAVs have a strong safety profile, recent studies show that AAV causes dendritic loss and synaptic weakening in mouse somatosensory cortex, impacts that are prevented by systemic administration of blockers of Toll-like receptor 9 (TLR9), an innate immunoreceptor that detects unmethylated cytosine-guanine (CpG) motifs. However, TLR9 blockers are immunosuppressive and costly. To realize AAVs full potential, it is critical to identify strategies to protect neurons without compromising immunity. Here we find that partially depleting CpG motifs from the AAV genome prevents AAV-induced dendritic loss and synaptic weakening. CpG depletion of transgene and intronic regions via codon-optimized synonymous mutations was protective and did not impair transgene expression in vivo. To facilitate the production and use of lower-CpG AAVs, we created a web-facing application, CpG-Assist Tool (CpG-AT), which allows researchers to quantify the CpG content of any sequence, explore the CpG content of commonly used AAV components, and generate CpG-depleted coding and non-coding sequences. This study identifies CpG depletion as a practical strategy to prevent AAV-induced neural circuit disruption, and provides tools to facilitate CpG reduction to enhance the safety and efficacy of AAV-mediated gene delivery.

neuroscience↗

bioRxiv2023

Tesofensine, a novel antiobesity drug, silences GABAergic hypothalamic neurons

Obesity is a major global health epidemic that has adverse effects on both the people affected as well as the cost to society. Several anti-obesity drugs that target GLP-1 receptors have recently come to the market. Here we describe the effects of tesofensine, a novel anti-obesity drug that acts as a triple monoamine neurotransmitter reuptake inhibitor. We investigated its effects on weight loss and underlying neuronal mechanisms in mice and rats using various techniques. These include behavioral tasks, DeepLabCut videotaped analysis, electrophysiological ensemble recordings, optogenetic activation, and chemogenetic silencing of GABAergic neurons in the Lateral Hypothalamus (LH). We found that tesofensine induced greater weight loss in obese than lean rats, which was associated with changes in LH ensemble activity. In Vgat-ChR2 and Vgat-IRES-cre transgenic mice, we found for the first time that tesofensine inhibited a subset of LH GABAergic neurons, reducing their ability to promote feeding behavior, and chemogenetically silencing them enhanced tesofensines food-suppressing effects. Unlike phentermine, a dopaminergic appetite suppressant, tesofensine causes few, if any, head-weaving stereotypy at therapeutic doses. Most importantly, we found that tesofensine prolonged the weight loss induced by 5-HTP, a serotonin precursor, and blocked the body weight rebound that often occurs after weight loss. Behavioral studies on rats with the tastant sucrose indicated that tesofensines appetite suppressant effects are independent of taste aversion and do not directly affect the perception of sweetness or palatability of sucrose. In summary, our data provide new insights into the effects of tesofensine on weight loss and the underlying neuronal mechanisms, suggesting that tesofensine may be an effective treatment for obesity and that it may be a valuable adjunct to other appetite suppressants to prevent body weight rebound.

neuroscience↗