Prenatal alcohol exposure potentiates chronic neuropathic pain, spinal glial and immune cell activation and alters sciatic nerve and DRG cytokine levels

Abstract

A growing body of evidence indicates that prenatal alcohol exposure (PAE) may predispose individuals to secondary medical disabilities later in life. Animal models of PAE reveal neuroimmune sequelae such as elevated brain astrocyte and microglial activation with corresponding region-specific changes in immune signaling molecules such as cytokines and chemokines. The aim of this study was to evaluate the effects of moderate PAE on the development and maintenance of allodynia induced by chronic constriction injury (CCI) of the sciatic nerve in adult male rat offspring. Because CCI allodynia requires the actions of glial cytokines, we analyzed lumbar spinal cord glial and immune cell surface markers indicative of their activated levels, as well as sciatic nerve and dorsal root ganglia (DRG) cytokines in PAE offspring in adulthood. While PAE did not alter basal sensory thresholds before or after sham manipulations, PAE significantly potentiated adult onset and maintenance of allodynia. Microscopic analysis revealed exaggerated astrocyte and microglial activation, while flow cytometry data demonstrated increased proportions of immune cells with cell surface major histocompatibility complex II (MHCII) and β-integrin adhesion molecules, which are indicative of PAE-induced immune cell activation. Sciatic nerves from CCI rats revealed that PAE potentiated the proinflammatory cytokines interleukin (IL)-1β, IL-6 and tumor necrosis factor-alpha (TNFα) protein levels with a simultaneous robust suppression of the anti-inflammatory cytokine, IL-10. A profound reduction in IL-10 expression in the DRG of PAE neuropathic rats was also observed. Taken together, our results provide novel insights into the vulnerability that PAE produces for adult-onset central nervous system (CNS) pathological conditions from peripheral nerve injury.

Cytokines in Pain: Harnessing Endogenous Anti-Inflammatory Signaling for Improved Pain Management

“…The role of LFA-1 in pathological pain was first explored in work by Noor et al that revealed prenatal alcohol exposure (PAE) predisposes adult rats to develop neuropathic pain in response to a minor injury. They found that LFA-1 expression is increased on microglia and trafficking immune cells in the lumbar spinal cords and on peripheral immune cells of neuropathic PAE rats (192). It was later identified that intrathecal or intravenous administration of BIRT-377, a small molecule non-competitive LFA-1 antagonist, relieves allodynia in neuropathic rats, which was associated with decreased immunoreactivity for spinal markers of glial activation and IL-1β and concurrent increases in IL-10 immunoreactivity (193,194).…”

LFA-1 antagonist (BIRT377) similarly reverses peripheral neuropathic pain in male and female mice with underlying sex divergent peripheral immune proinflammatory phenotypes

“…The total number of positive and negative responses were then entered into the computer software program, PsychoFit (http://psych.colorado.edu/~lharvey: RRID: SCR_015381) to determine the absolute withdrawal threshold (50% paw withdrawal threshold), as previously described [50] . The PsychoFit program fits a Gaussian integral psychometric function to the observed withdrawal rates for each monofilament using a maximum-likelihood fitting method [47,51] . The interpolated 50% withdrawal thresholds were then used for statistical analysis.…”

“…Tissue collection was conducted in six cohorts of mice (8 mice in each cohort, N of 1 from each experimental condition) as previously described [47,51] and modified as described here. Immediately following behavioral analysis on Day 13 post-surgery (Day 3 post-injection), mice were deeply anesthetized under isoflurane (10 min in 5% isoflurane and in oxygen at 2 L/min), followed by rapid transcardial perfusion with ice cold 0.1 M phosphate buffered saline (PBS; pH = 7.4; flow rate 10 mL/min).…”

“…Cellular lysates (supernatant) were collected in a new set of tubes, and protein concentrations were measured by Quickstart TM Bradford Protein Assay Kit (Biorad, CA, USA). Splenic cytokine and chemokine levels were determined using V-Plex TM multiplex immunoassays (MesoScale Discovery), as described previously [47,51,62,63] . Briefly, calibrators (provided by the kit) or samples (100 μg protein from each experimental sample per well) were loaded onto a "multi-spot" plate in duplicates.…”

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“…The centrally acting drugs (opiate analgesics) such as narcotics inhibited both phases equally, while antiinflammatory drugs (non-opiate analgesics) specially inhibited the second phase 20,21 . As we mentioned before it is also well known that the formalin model may involve sensorial C-fibers 22 in early phase and a combined process generated by peripheral inflammatory tissue and functional changes in the dorsal horn in late phase 23,24 .…”