An opioid receptor-independent mechanism underlies motility dysfunction and visceral hyperalgesia in opioid-induced bowel dysfunction.

Constipation and abdominal pain are commonly encountered in opioid-induced bowel dysfunction (OBD). The underlying mechanisms are incompletely understood, and treatments are not satisfactory. As patients with OBD often have fecal retention, we aimed to determine whether fecal retention plays a pathogenic role in the development of constipation and abdominal pain in OBD, and if so to investigate the mechanisms. A rodent model of OBD was established by daily morphine treatment at 10 mg/kg for 7 days. Bowel movements, colonic muscle contractility, visceromotor response to colorectal distention, and cell excitability of colon-projecting dorsal root ganglion neurons were determined in rats fed with normal pellet food, or with clear liquid diet. Morphine treatment (Mor) reduced fecal outputs starting on day 1, and caused fecal retention afterward. Compared with controls, Mor rats demonstrated suppressed muscle contractility, increased neuronal excitability, and visceral hypersensitivity. Expression of cyclooxygenase-2 (COX-2) and nerve growth factor (NGF) was upregulated in the smooth muscle of the distended colon in Mor rats. However, prevention of fecal retention by feeding rats with clear liquid diet blocked upregulation of COX-2 and NGF, restored muscle contractility, and attenuated visceral hypersensitivity in Mor rats. Moreover, inhibition of COX-2 improved smooth muscle function and fecal outputs, whereas anti-NGF antibody administration attenuated visceral hypersensitivity in Mor rats. Morphine-induced fecal retention is an independent pathogenic factor for motility dysfunction and visceral hypersensitivity in rats with OBD. Liquid diet may have therapeutic potential for OBD by preventing fecal retention-induced mechanotranscription of COX-2 and NGF.NEW & NOTEWORTHY Our preclinical study shows that fecal retention is a pathogenic factor in opioid-induced bowel dysfunction, as prevention of fecal retention with liquid diet improved motility and attenuated visceral hyperalgesia in morphine-treated animals by blocking expression of cyclooxygenase-2 and nerve growth factor in the colon.

Efficacy, Safety, and Feasibility of the Morphine Microdose Method in Community-Based Clinics.

Objectives: The goal of this study was to assess the success of the morphine microdose method in a community pain clinic setting by monitoring follow-up frequency, dose escalation, and monotherapy/polytherapy ratio. The morphine microdose method involves a pretrial reduction or elimination of systemic opioids followed by a period of abstinence. Intrathecal (IT) morphine is then started at doses of less than 0.2 mg per day. Systemic opioid abstinence is then continued after pump implant and IT morphine monotherapy. Design: Retrospective review of medical records. Setting: Private and academic pain clinic practices. Subjects: Chronic noncancer pain patients. Methods: We reviewed the charts of 60 patients who had completed a microdose regimen and had an IT pump implanted between June 11, 2008, and October 11, 2014. During IT therapy, dose change over time, pain scores, side effects, max dose, and duration were recorded. Results: The majority of patients (35/60, 58%) were successfully managed solely on morphine microdose monotherapy. These patients did not require additional oral therapy. There was a significant reduction in mean pain scores, from 7.4 ± 0.32 before microdose therapy to 4.8 ± 0.3 after microdose therapy. Conclusions: Microdose therapy achieved analgesia, improved safety, and avoided systemic side effects. The safety of IT therapy was increased by using a lower concentration (2 mg/mL) and lower daily doses (<3 mg/d) of morphine. Furthermore, microdose therapy was feasible, safe, and cost-effective in the outpatient setting.

A simple and fast method to image calcium activity of neurons from intact dorsal root ganglia using fluorescent chemical Ca2+ indicators.

Chemical calcium indicators have been commonly used to monitor calcium (Ca2+) activity in cell bodies, i.e., somata, of isolated dorsal root ganglion neurons. Recent studies have shown that dorsal root ganglion somata play an essential role in soma-glia interactions and actively participate in the transmission of nociceptive signals. It is therefore desirable to develop methods to study Ca2+ activity in neurons and glia in intact dorsal root ganglia. In our previous studies, we found that incubation of intact dorsal root ganglia with acetoxymethyl dye resulted in efficient Ca2+ dye loading into glial cells but limited dye loading into neurons. Here, we introduce a useful method to load Ca2+ dyes in intact dorsal root ganglion neurons through electroporation. We found that electroporation greatly facilitated loading of Fluo-4 acetoxymethyl, Oregon green bapta-1-488 acetoxymethyl, and Fluo-4 pentapotassium salt into dorsal root ganglion neurons. In contrast, electroporation did not further facilitate dye loading into glia. Using electroporation followed by incubation of acetoxymethyl form Ca2+ dye, we can load acetoxymethyl Ca2+ dye well in both neurons and glia. With this approach, we found that inflammation induced by complete Freund's adjuvant significantly increased the incidence of neuron-glia interactions in dorsal root ganglia. We also confirmed the actions of capsaicin and morphine on Ca2+ responses in dorsal root ganglion neurons. Thus, by promoting the loading of Ca2+ dye in neurons and glia through electroporation and incubation, Ca2+ activities in neurons and neuron-glia interactions can be well studied in intact dorsal root ganglia.

Pathogenesis of abdominal pain in bowel obstruction: role of mechanical stress-induced upregulation of nerve growth factor in gut smooth muscle cells.

Abdominal pain is one of the major symptoms in bowel obstruction (BO); its cellular mechanisms remain incompletely understood. We tested the hypothesis that mechanical stress in obstruction upregulates expression of nociception mediator nerve growth factor (NGF) in gut smooth muscle cells (SMCs), and NGF sensitizes primary sensory nerve to contribute to pain in BO. Partial colon obstruction was induced with a silicon band implanted in the distal bowel of Sprague-Dawley rats. Colon-projecting sensory neurons in the dorsal root ganglia (T13 to L2) were identified for patch-clamp and gene expression studies. Referred visceral sensitivity was assessed by measuring withdrawal response to stimulation by von Frey filaments in the lower abdomen. Membrane excitability of colon-projecting dorsal root ganglia neurons was significantly enhanced, and the withdrawal response to von Frey filament stimulation markedly increased in BO rats. The expression of NGF mRNA and protein was increased in a time-dependent manner (day 1-day 7) in colonic SMC but not in mucosa/submucosa of the obstructed colon. Mechanical stretch in vitro caused robust NGF mRNA and protein expression in colonic SMC. Treatment with anti-NGF antibody attenuated colon neuron hyperexcitability and referred hypersensitivity in BO rats. Obstruction led to significant increases of tetrodotoxin-resistant Na currents and mRNA expression of Nav1.8 but not Nav1.6 and Nav1.7 in colon neurons; these changes were abolished by anti-NGF treatment. In conclusion, mechanical stress-induced upregulation of NGF in colon SMC underlies the visceral hypersensitivity in BO through increased gene expression and activity of tetrodotoxin-resistant Na channels in sensory neurons.

Communication between neuronal somata and satellite glial cells in sensory ganglia.

Studies of the structural organization and functions of the cell body of a neuron (soma) and its surrounding satellite glial cells (SGCs) in sensory ganglia have led to the realization that SGCs actively participate in the information processing of sensory signals from afferent terminals to the spinal cord. SGCs use a variety ways to communicate with each other and with their enwrapped soma. Changes in this communication under injurious conditions often lead to abnormal pain conditions. "What are the mechanisms underlying the neuronal soma and SGC communication in sensory ganglia?" and "how do tissue or nerve injuries affect the communication?" are the main questions addressed in this review.

Region-specific generation of cholinergic neurons from fetal human neural stem cells grafted in adult rat.

Pluripotent or multipotent stem cells isolated from human embryos or adult central nervous system (CNS) may provide new neurons to ameliorate neural disorders. A major obstacle, however, is that the majority of such cells do not differentiate into neurons when grafted into non-neurogenic areas of the adult CNS. Here we report a new in vitro priming procedure that generates a nearly pure population of neurons from fetal human neural stem cells (hNSCs) transplanted into adult rat CNS. Furthermore, the grafted cells differentiated by acquiring a cholinergic phenotype in a region-specific manner. This technology may advance stem cell-based therapy to replace lost neurons in neural injury or neurodegenerative disorders.