Inhibition of p75 Neurotrophin Receptor Attenuates Isoflurane-mediated Neuronal Apoptosis in the Neonatal Central Nervous System

Background. Exposure to anesthetics during synaptogenesis results in apoptosis and subsequent cognitive dysfunction in adulthood. Probrain-derived neurotrophic factor (proBDNF) Is involved in synaptogenesis and can induce neuronal apoptosis via p75 neurotrophic receptors (p75(NTR)). proBDNF is cleaved into mature BDNF (mBDNF) by plasmin, a protease converted from plasminogen by tissue plasminogen activator (tPA) that is released with neuronal activity; mBDNF supports survival and stabilizes synapses through tropomyosin receptor kinase R. The authors hypothesized that anesthetics suppress tPA release from neurons, enhance p75(NTR) signaling, and reduce synapses, resulting in apoptosis. Methods: Primary neurons (DIV5) and postnatal day 5-7 (PND5-7) mice were exposed to Isoflurane 0.4%, 4 h) in 5% CO2, 95% air. Apoptosis was assessed by cleaved caspase-3 (Cl-Csp3) Immunoblot and immunofluorescence microscopy. Dendritic spine changes were evaluated with the neuronal spine marker, drebrin. Changes in synapses in PNDS-7 mouse hippocampi were assessed by electron microscopy. Primary neurons were exposed to tPA, plasmin, or pharmacologic inhibitors of p75(NTR) (Fc-p75(NTR) or TAT-Pcp5) 15 min before isoflurane. TAT-Pcp5 was administered by intraperitoneal Injection to PND5-7 mice 15 min before isoflurane. Results: Exposure of neurons In vitro (DIV5) to isoflurane decreased tPA In the culture medium, reduced drebrin expression (marker of dendritic filopodial spines), and enhanced Cl-Csp3. tPA, plasmin, or TAT-Pcp5 stabilized dendritic filopodial spines and decreased Cl-Csp3 in neurons. TAT-Pep5 blocked isoflurane-mediated Increase in Cl-Csp3 and reduced In PND5-7 mouse hippocampi. Conclusion: tPA, plasmin, or p75(NTR) inhibition blocked isoflurane-mediated reduction in dendritic filopodial spines and neuronal apoptosis in vitro. Isoflurane reduced synapses and enhanced Cl-Csp3 in the hippocampus of PND5-7 mice, the latter effect being mitigated by p75(NTR) inhibition in vivo. These data support the hypothesis that isoflurane neurotoxicity in the developing rodent brain is mediated by reduced synaptic tPA release and enhanced proBDNF/p75(NTR)-mediated apoptosis.