Neurite Degeneration Induced by Heme Deficiency Mediated via Inhibition of NMDA Receptor-Dependent Extracellular Signal-Regulated Kinase 1/2 Activation

Abstract

The early stages of many neurodegenerative diseases and age-related degeneration are characterized by neurite damage and compromised synaptic function that precede neuronal cell death. We investigated the signaling mechanisms underlying neurite degeneration using cortical neuron cultures. Inhibition of heme synthesis caused neurite damage, without neuronal death, and was mediated by reduced NMDA receptor (NMDAR) expression and phosphorylation. The signaling toward the degenerative phenotype involved suppression of the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway, and electrophysiological recording showed that the neurodegeneration is accompanied by reduced NMDAR current and Ca 2ϩ influx, as well as reduced voltage-gated sodium currents, consistent with compromised neurite integrity. Rescue from the degenerative phenotype by heme replacement was dependent on restoration of NR2B subunit phosphorylation and expression of NMDAR currents with higher Ca 2ϩ permeability, consistent with triggering prosurvival ERK1/2 signaling to maintain and extend neurites. This study demonstrated a new mechanism of neurodegeneration in which impaired heme synthesis led to NMDAR signaling dysfunction, suppression of the prosurvival ERK1/2 pathway, and progressive fragmentation of neuronal projections.

Early Failure ofN-Methyl-d-aspartate Receptors and Deficient Spine Formation Induced by Reduction of Regulatory Heme in Neurons

“…Heme-deficient cultured neurons displayed progressive neurite fragmentation followed, eventually, by cell death. The rescue of prosurvival extracellular signal-regulated kinase 1/2 activation by heme was mediated predominantly via the NR2B-containing NMDARs (Chernova et al, 2007). These observations were made in neurons chronically deficient in heme, in which the loss of both bound and regulatory heme pools might have contributed to the progressive fragmentation of neurites.…”

“…For measurement of heme synthesis, cells were incubated with 0.4 Ci of [3,5-3 H]aminolevulinic acid hydrochloride (2.6 Ci/mmol; PerkinElmer Life and Analytical Sciences, Waltham, MA) for 24 h. Heme was extracted from the cells by acetone-HCl and diethyl ether. The amount of radioactivity in extracted heme was measured by scintillation counting as described previously (Chernova et al, 2007). Total recovery of radioactivity from all fractions was the same for treated and untreated cells.…”

“…Although continuously hemedeficient neurons have reduced expression of NMDAR subunits at the stage of neurite loss (Chernova et al, 2007), the initial changes after inhibition of heme synthesis do not involve down-regulation in gene or protein expression for NMDAR subunits NR1 and NR2B (Supplemental Fig. 1, A-D).…”

“…1, A-D). NR2B is a major tyrosine-phosphorylated protein regulated by SFKs in the postsynaptic density (Moon et al, 1994) and is impaired in heme-deficient neurons (Chernova et al, 2007). To examine the early changes caused by reduced heme availability, we used immunoblotting and immunostaining to monitor NR2B phosphorylation at Tyr1336 and Tyr1252 in the presence or absence of SFKs inhibitor PP2 and heme in neurons treated with SA for 24 h, the time point at which the 2-fold up-regulation of ALAS1 indicated a depletion of the regulatory heme pool (Fig.…”

B2 SINE retrotransposon causes polymorphic expression of mouse 5-aminolevulinic acid synthase 1 gene

“…The ability of heme to bind gases reversibly accounts for its functions in other hemoproteins in transporting oxygen, and as a sensor of O 2, NO and CO [2]. Heme also has many roles in the control of gene expression and cellular metabolism as a signalling molecule modulating the activity of ion channels, transcription factors and kinases [3,4], microRNA processing [5] and the N-rule pathway of protein degradation [6].…”