Abstracts

Rationale: Children born preterm are prone to chronic neurological deficits including cognitive delay, epilepsy, and behavioral abnormalities. Alterations in initial inhibitory circuit development and changes in inhibitory tone have been implicated in the etiology of these disorders. The primary inhibitory neurotransmitter in the adult brain is GABA. As the cerebral cortex matures and responds to environmental cues during development, GABA responses switch from excitatory to inhibitory coincident with upregulation of the potassium chloride cotransporter KCC2. Developmental increases in KCC2 allows GABA receptor activation to cause hyperpolarization and aids in attenuating hyperexcitability. Previously we showed decreased cortical KCC2 protein expression in preterm infants with white matter lesions and in patients undergoing epilepsy resections. Here, we hypothesize a transient systemic hypoxic-ischemic (TSHI) insult to the developing brain during late gestation alters the spatiotemporal pattern of KCC2 upregulation, thereby disrupting essential cortical inhibitory circuit development. Methods: Bilateral uterine artery ligation was performed on E18 rats to induce TSHI typical in early preterm birth with placental insufficiency. Erythropoietin (EPO) or vehicle (Veh) was administered following TSHI from P1-5 (2000 IU/kg/dose/ip). Changes in NKCC1 and KCC2 protein were investigated and whole-cell voltage clamp of CA3 pyramidal neurons was used to determine miniature inhibitory postsynaptic currents (mIPSCs) and assess inhibitory synaptic input. Results: Results demonstrate a developmental upregulation of KCC2 protein in CA3 from P7-21, concomitant with a decreased NKCC1 to KCC2 ratio consistent with previous reports. Following TSHI, KCC2 protein expression was significantly reduced at P13 by 62% compared to sham. Postnatal administration of EPO attenuated the reduction in KCC2 protein expression (TSHI+Veh: 39% control vs. TSHI+EPO: 155% control).TSHI (n=12 cells/7 rats) also decreased the mean amplitude, frequency and rise time of mIPSCs at P10-11, compared to sham (n=11 cells/6 rats). Two distinct populations of TSHI CA3 pyramidal neurons emerged from mIPSC frequency analysis, with 67% of cells showing a frequency <0.3 Hz, compared to 0% of sham cells (p<0.001). EPO normalized the effects of TSHI and restored amplitude and frequency to baseline levels. Conclusions: These results indicate decreased inhibitory synaptic input and KCC2 protein expression following prenatal HI, and may have important implications for understanding inhibitory circuit development and the effect of late gestation insults on inhibitory tone. Importantly, the postnatal administration of EPO normalized TSHI-induced reductions in KCC2 protein expression and mIPSC amplitude and frequency, suggesting it is possible to reverse abnormalities in inhibitory cortical development. Together, these data indicate that the impairments caused by prenatal insults in children born preterm, including cognitive delay and epilepsy, may be at least partially restored with a neuroprotective regimen administered in the neonatal period.