Early Postnatal Exposure to Isoflurane Disrupts Oligodendrocyte Development and Myelin Formation in the Mouse Hippocampus

Abstract

Editor’s Perspective What We Already Know about This Topic What This Article Tells Us That Is New Background Early postnatal exposure to general anesthetics may interfere with brain development. We tested the hypothesis that isoflurane causes a lasting disruption in myelin development via actions on the mammalian target of rapamycin pathway. Methods Mice were exposed to 1.5% isoflurane for 4 h at postnatal day 7. The mammalian target of rapamycin inhibitor, rapamycin, or the promyelination drug, clemastine, were administered on days 21 to 35. Mice underwent Y-maze and novel object position recognition tests (n = 12 per group) on days 56 to 62 or were euthanized for either immunohistochemistry (n = 8 per group) or Western blotting (n = 8 per group) at day 35 or were euthanized for electron microscopy at day 63. Results Isoflurane exposure increased the percentage of phospho-S6–positive oligodendrocytes in fimbria of hippocampus from 22 ± 7% to 51 ± 6% (P < 0.0001). In Y-maze testing, isoflurane-exposed mice did not discriminate normally between old and novel arms, spending equal time in both (50 ± 5% old:50 ± 5% novel; P = 0.999), indicating impaired spatial learning. Treatment with clemastine restored discrimination, as evidenced by increased time spent in the novel arm (43 ± 6% old:57 ± 6% novel; P < 0.001), and rapamycin had a similar effect (44 ± 8% old:56 ± 8% novel; P < 0.001). Electron microscopy shows a reduction in myelin thickness as measured by an increase in g-ratio from 0.76 ± 0.06 for controls to 0.79 ± 0.06 for the isoflurane group (P < 0.001). Isoflurane exposure followed by rapamycin treatment resulted in a g-ratio (0.75 ± 0.05) that did not differ significantly from the control value (P = 0.426). Immunohistochemistry and Western blotting show that isoflurane acts on oligodendrocyte precursor cells to inhibit both proliferation and differentiation. DNA methylation and expression of a DNA methyl transferase 1 are reduced in oligodendrocyte precursor cells after isoflurane treatment. Effects of isoflurane on oligodendrocyte precursor cells were abolished by treatment with rapamycin. Conclusions Early postnatal exposure to isoflurane in mice causes lasting disruptions of oligodendrocyte development in the hippocampus via actions on the mammalian target of rapamycin pathway.

General Anesthetic-Induced Neurotoxicity in the Immature Brain: Reevaluating the Confounding Factors in the Preclinical Studies

“…As shown in rodents, neonatal exposure to sevoflurane, isoflurane, nitric oxide, propofol, ketamine, and others was found to induce apoptosis of neurons and oligodendrocytes and long-term cognitive impairment in rhesus monkeys [10,[14][15][16][17][18][19][20][21][22]48]. Notably, a new important advance in this field was that recent findings showed that GA disrupted myelin formation in the developing brain of NHPs and mice [49,50]. Zhang et al's report was the first finding that sevoflurane exposure in early life disrupted folate metabolism and then interrupted oligodendrocyte development in NHPs [50].…”

Effects of Early Exposure of Isoflurane on Chronic Pain via the Mammalian Target of Rapamycin Signal Pathway

“…The mice were exposed to 1.5% isoflurane carried with CG for 4 h. A calibrated flowmeter was used to deliver CG at a flow rate of 5 L/min, and an agent-specific vaporizer was used to deliver isoflurane. After isoflurane exposure, mice were returned to their dams together with their littermates upon regaining righting reflex [51].…”

“…Vehicle consisted of 5% Tween 80 (Sigma-Aldrich, St. Louis, MO, USA), 10% polyethylene glycol 400 (Sigma-Aldrich, St. Louis, MO, USA), and 8% ethanol in saline. No treatment was used for control animals (group 1) [51].…”

“…All c-fos-positive (labeled in nuclei) and pS6-immunolabeled neurons in IC, ACC, and superficial SDH (lamina I and II) were counted and quantitatively analyzed using the ImageJ program (NIH, v.1.52p, Bethesda, MD, USA). Double-labeled cells were identified in channel-merged images [51].…”

“…Images were acquired using ChemiDoc imaging system (BioRad, Hercules, CA, USA) and were quantitated with ImageJ (NIH, v.1.52p, Bethesda, MD, USA). The ratios of band density from p-mTOR over t-mTOR or PSD95 over β-actin were calculated, respectively, and quantitative analysis was performed with the ImageJ program [51]. Each band that is shown in the figures represents a single trial with a unique animal.…”