Peripheral neuropathy is a frequent complication of chronic diabetes that most commonly presents as a distal degenerative polyneuropathy with sensory loss. Around 20% to 30% of such patients may also experience neuropathic pain. The underlying pathogenic mechanisms are uncertain, and therapeutic options are limited. Rodent models of diabetes have been used for more than 40 years to study neuropathy and evaluate potential therapies. For much of this period, streptozotocin-diabetic rats were the model of choice. The emergence of new technologies that allow relatively cheap and routine manipulations of the mouse genome has prompted increased use of mouse models of diabetes to study neuropathy. In this article, we describe the commonly used mouse models of type 1 and type 2 diabetes, and provide protocols to phenotype the structural, functional, and behavioral indices of peripheral neuropathy, with a particular emphasis on assays pertinent to the human condition. © 2016 by John Wiley & Sons, Inc.

Preclinical studies indicate that diverse muscarinic receptor antagonists, acting via the M₁ sub-type, promote neuritogenesis from sensory neurons in vitro and prevent and/or reverse both structural and functional indices of neuropathy in rodent models of diabetes. We sought to translate this as a potential therapeutic approach against structural and functional indices of diabetic neuropathy using oxybutynin, a muscarinic antagonist approved for clinical use against overactive bladder. Studies were performed using sensory neurons maintained in vitro, rodent models of type 1 or type 2 diabetes and human subjects with type 2 diabetes and confirmed neuropathy. Oxybutynin promoted significant neurite outgrowth in sensory neuron cultures derived from adult normal rats and STZ-diabetic mice, with maximal efficacy in the 1-100 nmol/l range. This was accompanied by a significantly enhanced mitochondrial energetic profile as reflected by increased basal and maximal respiration and spare respiratory capacity. Systemic (3-10 mg/kg/day s.c.) and topical (3% gel daily) oxybutynin reversed paw heat hypoalgesia in the STZ and db/db mouse models of diabetes and reversed paw tactile allodynia in STZ-diabetic rats. Loss of nerve profiles in the skin and cornea of db/db mice was also prevented by daily topical delivery of 3% oxybutynin for 8 weeks. A randomized, double-blind, placebo-controlled interventional trial was performed in subjects with type 2 diabetes and established peripheral neuropathy. Subjects received daily topical treatment with 3% oxybutynin gel or placebo for 6 months. The a priori designated primary endpoint, significant change in intra-epidermal nerve fibre density (IENFD) in skin biopsies taken before and after 20 weeks of treatments, was met by oxybutynin but not placebo. Secondary endpoints showing significant improvement with oxybutynin treatment included scores on clinical neuropathy, pain and quality of life scales. This proof-of-concept study indicates that muscarinic antagonists suitable for long-term use may offer a novel therapeutic opportunity for treatment of diabetic neuropathy. Trial registry number: NCT03050827.