Biology subjects

Mwirigi, J.

Publications and source records attributed to Mwirigi, J..

4 recordsLinked to original sources

bioRxiv2026

Preferential innervation of endometriosis by hyperexcitable Ret/GFRα1+ nociceptors associates with target GDNF and clinical pain

Endometriosis is a prevalent condition characterized by chronic pelvic pain that is frequently refractory to treatment. While the mechanisms underlying this pain remain poorly defined, clinical evidence often indicates that lesion innervation, but not disease stage (e.g. number and depth of lesions), correlate with pelvic pain severity. However, characterization of lesion-innervating neurons is incomplete, revealing an opportunity to identify novel, disease-modifying therapeutics. Here, we coupled functional analyses of lesion-innervating neurons in a mouse model with concurrent identification and characterization of lesion-innervating neurons from pain-phenotyped endometriosis patients. Following the confirmation of abdominal-directed pain-like behaviors in the mouse model, electrophysiological analysis revealed that lesion-innervating dorsal root ganglion (DRG) neurons are hyperexcitable compared to matched controls. These neurons are predominantly small-diameter and bind Isolectin B4, an established marker of the GDNF Family Ligand receptor, Ret. GDNF is concentrated within the stromal layer of both mouse and human lesions, adjacent to axons expressing the GDNF co-receptor, GFR1. Critically, clinical pain correlates with lesion GDNF level, axonal density, and neuronal GFR1 levels. These data provide evidence that endometrial lesions may recruit the Ret-positive subpopulation of nociceptors where they become sensitized and increase patient pain.

neuroscience↗

bioRxiv2026

The Critical Role of Pdyn-Lineage Enteric Neurons in Colonic Motility and Visceral Interoception

Summary/Abstract Sensory neurons play well defined roles in the regulation of intestinal motility, digestion, and interoception, but transcriptional dissection of intrinsic and extrinsic sensory neurons innervating the intestines has been challenging because these cells share many genetic markers. However, we cross-referenced transcriptional profiles of intrinsic and extrinsic intestinal neurons and the cells that surround them and identified Pdyn, the gene encoding Prodynorphin, as a marker of putative sensory enteric neurons of the mouse intestines. In a Pdyn lineage-reporter mouse, we identified labeled cells in the myenteric and submucosal plexuses of the large intestine, in contrast to their sparse presence in the dorsal root or nodose ganglia. In dissociated cell culture, these neurons mostly display a phasic firing pattern, discharging one or two action potentials at the onset of a depolarizing current pulse, followed by a prompt cessation of firing despite continued current injection, as assessed by whole-cell patch-clamp recordings. Optogenetic activation of Pdyn-lineage neurons propels stool in ex vivo colons, and in untethered and mobile mice, optogenetic stimulation of the proximal colon induces freezing behaviors and orbital tightening, suggesting interoceptive behaviors, without significant stool output differences. Together, these findings suggest that activation of Pdyn-lineage enteric neurons regulates motility and visceral interoception.

neuroscience↗

bioRxiv2025

Single nucleus multiomic atlas of human dorsal root ganglia reveals the contribution of non-neuronal cell types to pain

Sensory neurons residing in dorsal root ganglia (DRG) transmit sensory information such as pain, itch, touch, pressure and bodily position to the central nervous system. The activity of sensory neurons is regulated by non-neuronal cell types in the DRG, including satellite glial cells (SGCs), immune cells and fibroblasts. Dysregulated gene expression in DRG cells contributes to sensory nervous system disorders such as chronic pain. Understanding the genetic underpinnings of these conditions requires dissecting transcriptional regulation in human tissue. In this study, we profiled transcriptomic and chromatin accessibility landscapes from postmortem human DRG (hDRG) samples at single-nucleus level. We demonstrate that sequencing depth significantly impacts downstream analysis, with deeper sequencing yielding more detected cells and features, improved data integration, refined clustering and annotation, and more accurate scientific interpretations. We identified nine major cell types, defined their molecular signatures, and mapped cis-regulatory landscapes. Integration of gene expression with chromatin accessibility enabled peak-gene association and transcriptional network analyses, revealing transcription factors, their target genes, regulatory elements and potential partners that cooperatively drive cell-type-specific gene expression programs. This integrative approach identified cell types, genes, and cis-regulatory regions potentially driving pain conditions. Our unbiased genome-wide analysis not only recovered known pain-related genes but also highlighted novel candidate genes and regulatory regions implicated in pain mechanisms. Importantly, our results demonstrate that non-neuronal cells, including endothelial cells, fibroblasts, macrophages, and SGCs, play critical roles in pain pathogenesis and should be investigated as therapeutic targets. One Sentence SummaryOur work provides a comprehensive single-nucleus multi-omic atlas of human dorsal root ganglia, uncovering novel cell-type-specific regulatory mechanisms and candidate therapeutic targets for pain, thereby directly advancing translational insights into human sensory disorders and chronic pain pathogenesis.

neuroscience↗

bioRxiv2024

Genetic editing of primary human dorsal root ganglion neurons using CRISPR-Cas9 with functional confirmation

CRISPR-Cas9 editing is now the leading method for genome editing and is being advanced for the treatment of human disease. CRIPSR editing could have many applications for treatment of neurological diseases, including pain but traditional viral vector delivery approaches have neurotoxicity limiting their use. Overcoming these issues could open the door for genome editing treatments for diseases like intractable pain where the dorsal root ganglia (DRG) would be the desired target. To this end, we describe a simple method for viral-vector-independent transfection of primary human DRG (hDRG) neurons for CRISPR-Cas9 editing. As proof of principle, we edited TRPV1, NTSR2, and CACNA1E using a lipofection method with CRISPR-Cas9 plasmids containing reporter tags (GFP or mCherry). Transfection was successful as demonstrated by the expression of the reporters as early as two days in vitro. CRISPR-Cas9 editing was confirmed at the genome level with insertion and deletion detection system T7-endonuclease-I assay; protein level with immunocytochemistry and Western blot; and functional level through capsaicin-induced Ca2+ accumulation in a high-throughput compatible fluorescent imaging plate reader (FLIPR) system. This work establishes a reliable, target specific, non-viral CRISPR-Cas9-mediated genetic editing in primary human neurons with potential for future clinical application for intractable pain. TeaserWe describe a non-viral transfection method for CRISPR-Cas9 gene editing in human dorsal root ganglion neurons.

neuroscience↗