Mechanisms of neonatal erythropoietin neuroprotection

Robinson, Shenandoah

Abstract

Infants who are born very preterm are prone to cognitive delay, behavioral abnormalities, epilepsy and cerebral palsy. Medications and surgery can partially improve motor function and seizure control, but no interventions directly address the most common deficits - cognitive and behavioral problems, even though these impairments pose the biggest obstacles to these children becoming productive adults. Understanding the mechanisms of how early brain injury disrupts cerebral development, and moreover, how interventions restore cerebral function, will guide the indications and timing of promising therapies. Central nervous system (CNS) injury from very preterm birth often results from a global prenatal hypoxic-ischemic (HI) and/or inflammatory insult. Late in gestation subplate neurons guide and refine cerebral cortical circuit development, especially in cortical layer IV. Subplate neurons are essential for initiation of GABAergic inhibition in developing layer IV via spatiotemporal upregulation of the cation-chloride co-transporter KCC2 and maturation of GABAAR subunits. By extruding chloride, increasing KCC2 expression regulates the developmental switch of GABA responses from excitatory to inhibitory, and along with GABAAR maturation, directs effective cortical circuit formation. We hypothesize that CNS injury associated with preterm birth causes premature subplate neuron loss, and thus impairs maturation of cortical layer IV GABAAR subunits and functional KCC2 expression, essential components of cerebral cortical development. Further, we predict that post-injury neuroprotective erythropoietin (EPO) treatment can mitigate compromised cortical development by limiting alterations in GABAAR subunits and KCC2 maturation, and by promoting neurological recovery. Using our clinically-relevant model of prenatal transient systemic hypoxia-ischemia (TSHI) in rodents on embryonic day 18 (E18) that models the global CNS injury associated with very preterm birth in humans, we propose these Aims: 1) to test that premature loss of subplate following prenatal TSHI in rats impairs cerebral cortical maturation in vivo and in slice cultures with mechanical subplate ablation, 2) to test that prenatal TSHI limits KCC2 membrane expression in cortical layer IV via BDNF/calpain-mediated mechanisms, and 3) to test that post-injury neonatal EPO treatment promotes cortical layer IV KCC2 and GABAAR recovery after E18 TSHI in vivo and in vitro. We predict postnatal EPO treatment minimizes premature subplate regression, increases KCC2 expression and restores GABAAR subunit maturation. After prenatal TSHI followed by neonatal EPO or vehicle treatment, cognitive and behavioral function will be tested in young adult rats to test our prediction that EPO can restore cerebral cortical development. Together, these studies will elucidate mechanisms of subplate regulation of early cerebral cortical development following prenatal global HI, and provide insights into how prenatal injury is reversed with delayed EPO treatment, with the goal of improving cognitive and behavioral outcomes after prenatal injury.

Public Health Relevance

Infants who are born prematurely are prone to neurological problems including severe learning difficulties, and behavioral problems, such as autism, attention deficit disorder or anxiety. We propose that injury to the developing brain associated with preterm birth leads to long term consequences in brain function, and will define the molecular signaling mechanisms that mediate the neurological deficits. We also will test whether treatment with the neuroprotective agent erythropoietin can mitigate the damage to the developing brain.