Background Recent population studies have suggested that children with multiple exposures to anesthesia and surgery at an early age are at an increased risk of cognitive impairment. We therefore have established an animal model with multiple versus single exposures of anesthetic(s) in young versus adult mice, aiming to distinguish the role of different anesthesia in cognitive impairment. Methods Six day and 60 day-old mice were exposed to various anesthesia regimen. We then determined the effects of the anesthesia on learning and memory function, levels of pro-inflammatory cytokine interleukin-6 and tumor necrosis factor-α in brain tissues, and the amount of ionized calcium binding adaptor molecule 1 positive cells, the marker of microglia activation, in the hippocampus. Results Here we show that anesthesia with 3% sevoflurane two hours daily for three days induced cognitive impairment and neuroinflammation [e.g., increased interleukin-6 levels: 151% ± 2.3 (mean ± SD) versus 100% ± 9.0, P = 0.035, n = 6] in young, but not adult, mice. Anesthesia with 3% sevoflurane two hours daily for one day and 9% desflurane two hours daily for three days induced neither cognitive impairment nor neuroinflammation. Finally, an enriched environment and anti-inflammation treatment (ketorolac) ameliorated the sevoflurane anesthesia-induced cognitive impairment. Conclusions Anesthesia-induced cognitive impairment may depend on developmental stage, anesthetic agent, and the number of exposures. These findings also suggest the cellular basis and the potential prevention and treatment strategies for the anesthesia-induced cognitive impairment, which may ultimately lead to safer anesthesia care and better postoperative outcomes for children.

Diffuse white matter injury (DWMI), a leading cause of neurodevelopmental disabilities in preterm infants, is characterized by reduced oligodendrocyte formation. Oligodendrocyte precursor cells (NG2-cells) are exposed to various extrinsic regulatory signals, including the neurotransmitter GABA. We investigated GABAergic signaling to cerebellar white matter NG2-cells in a mouse model of DWMI (chronic neonatal hypoxia). We found that hypoxia caused a loss of GABAA receptor-mediated synaptic input to NG2-cells, extensive proliferation of these cells and delayed oligodendrocyte maturation, leading to dysmyelination. Treatment of control mice with a GABAA receptor antagonist or deletion of the chloride-accumulating transporter NKCC1 mimicked the effects of hypoxia. Conversely, blockade of GABA catabolism or GABA uptake reduced NG2-cell numbers and increased the formation of mature oligodendrocytes both in control and hypoxic mice. Our results indicate that GABAergic signaling regulates NG2-cell differentiation and proliferation in vivo, and suggest that its perturbation is a key factor in DWMI.

Na ϩ /Hϩ exchanger isoform 1 (NHE1) is a major acid extrusion mechanism after intracellular acidosis. We hypothesized that stimulation of NHE1 after cerebral ischemia contributes to the disruption of Na ϩ homeostasis and neuronal death. In the present study, expression of NHE1 was detected in cultured mouse cortical neurons. Three hours of oxygen and glucose deprivation (OGD) followed by 21 h of reoxygenation ( ϩ/ϩ mice. NHE1 ϩ/ϩ mice treated with HOE 642 or NHE1 heterozygous mice exhibited a ϳ33% decrease in infarct size ( p Ͻ 0.05). These results imply that NHE1 activity disrupts Na ϩ and Ca 2ϩ homeostasis and contributes to ischemic neuronal damage.

The brain achieves homeostasis of its intracellular and extracellular fluids by precisely regulating the transport of solute and water across its major cellular barriers: endothelia of the blood-brain barrier (BBB), choroid plexus epithelia, and neuroglial cell membranes. Cerebral edema, the pathological accumulation of fluid in the brain's intracellular and extracellular spaces, is a major cause of morbidity and mortality following stroke and other forms of ischemic brain injury. Until recently, mechanisms of cerebral edema formation have been obscure; consequently, its treatment has been empiric and suboptimal. Here, we provide a paradigm for understanding ischemic cerebral edema, showing that its molecular pathogenesis is a complex yet step-wise process that results largely from impaired astrocytic cell volume regulation and permeability alterations in the cerebral microvasculature, both of which arise from pathological changes in the activities of specific ion channels and transporters. Recent data has implicated the bumetanide-sensitive NKCC1, an electroneutral cotransporter expressed in astrocytes and the BBB, in cerebral edema formation in several different rodent models of stroke. Pharmacological inhibition or genetic deficiency of NKCC1 decreases ischemia-induced cell swelling, BBB breakdown, cerebral edema, and neurotoxicity. Combination pharmacological strategies that include NKCC1 as a target might thus prove beneficial for the treatment of ischemic, and potentially other types of, cerebral edema.

This review presents a brief overview of the γ-aminobutyric acid (GABA) system in the developing and mature central nervous system (CNS) and its potential connections to pathologies of the CNS. γ-aminobutyric acid (GABA) is a major neurotransmitter expressed from the embryonic stage and throughout life. At an early developmental stage, GABA acts in an excitatory manner and is implicated in many processes of neurogenesis, including neuronal proliferation, migration, differentiation, and preliminary circuit-building, as well as the development of critical periods. In the mature CNS, GABA acts in an inhibitory manner, a switch mediated by chloride/cation transporter expression and summarized in this review. GABA also plays a role in the development of interstitial neurons of the white matter, as well as in oligodendrocyte development. Although the underlying cellular mechanisms are not yet well understood, we present current findings for the role of GABA in neurological diseases with characteristic white matter abnormalities, including anoxic-ischemic injury, periventricular leukomalacia, and schizophrenia. Development abnormalities of the GABAergic system appear particularly relevant in the etiology of schizophrenia. This review also covers the potential role of GABA in mature brain injury, namely transient ischemia, stroke, and traumatic brain injury/post-traumatic epilepsy.

We reported previously that inhibition of Na(+)-K(+)-Cl(-) cotransporter isoform 1 (NKCC1) by bumetanide abolishes high extracellular K(+) concentration ([K(+)](o))-induced swelling and intracellular Cl(-) accumulation in rat cortical astrocytes. In this report, we extended our study by using cortical astrocytes from NKCC1-deficient (NKCC1(-/-)) mice. NKCC1 protein and activity were absent in NKCC1(-/-) astrocytes. [K(+)](o) of 75 mM increased NKCC1 activity approximately fourfold in NKCC1(+/+) cells (P < 0.05) but had no effect in NKCC1(-/-) astrocytes. Intracellular Cl(-) was increased by 70% in NKCC1(+/+) astrocytes under 75 mM [K(+)](o) (P < 0.05) but remained unchanged in NKCC1(-/-) astrocytes. Baseline intracellular Na(+) concentration ([Na(+)](i)) in NKCC1(+/+) astrocytes was 19.0 +/- 0.5 mM, compared with 16.9 +/- 0.3 mM [Na(+)](i) in NKCC1(-/-) astrocytes (P < 0.05). Relative cell volume of NKCC1(+/+) astrocytes increased by 13 +/- 2% in 75 mM [K(+)](o), compared with a value of 1.0 +/- 0.5% in NKCC1(-/-) astrocytes (P < 0.05). Regulatory volume increase after hypertonic shrinkage was completely impaired in NKCC1(-/-) astrocytes. High-[K(+)](o)-induced (14)C-labeled D-aspartate release was reduced by approximately 30% in NKCC1(-/-) astrocytes. Our study suggests that stimulation of NKCC1 is required for high-[K(+)](o)-induced swelling, which contributes to glutamate release from astrocytes under high [K(+)](o).

Na-K-Cl cotransporter isoform 1 (NKCC1) plays an important role in maintenance of intracellular Naϩ , K ϩ , and Cl Ϫ levels in astrocytes. We propose that NKCC1 may contribute to perturbations of ionic homeostasis in astrocytes under ischemic conditions. After 3-8 hr of oxygen and glucose deprivation (OGD), NKCC1-mediated 86 Rb influx was significantly increased in astrocytes from NKCC1 wild-type (NKCC1 ϩ/ϩ ) and heterozygous mutant (NKCC1 ϩ/Ϫ ) mice. Phosphorylated NKCC1 protein was increased in NKCC1 ϩ/ϩ astrocytes at 2 hr of OGD. Two hours of OGD and 1 hr of reoxygenation (OGD/REOX) triggered an ϳ3. IntroductionNa-K-Cl cotransporter isoform 1 (NKCC1) belongs to the cation-dependent Cl Ϫ transporter family and transports Na ϩ , K ϩ , and Cl Ϫ into cells under physiological conditions (Russell, 2000). NKCC1 is expressed in rat cortical astrocytes (Yan et al., 2001), oligodendrocytes (Hoppe and Kettenmann, 1989;Wang et al., 2003), and Schwann cells (Alvarez-Leefmans, 2001). Functions of NKCC1 in these cells include K ϩ and Na ϩ uptake (Su et al., 2000(Su et al., , 2002b and accumulation of Cl Ϫ above its electrochemical equilibrium (Hoppe and Kettenmann, 1989;Wang et al., 2003). In recent studies, we found that NKCC1 contributes to K ϩ uptake, swelling, and swelling-induced glutamate release in astrocytes in the presence of high extracellular K ϩ (Su et al., 2002a,b). Disruption of Naϩ and Ca 2ϩ homeostasis plays an important role in ischemic cell damage (Siesjö, 1992). A steep inwardly directed Na ϩ gradient is essential for glial functions, such as glutamate reuptake and regulation of intracellular ion concentrations by other secondary ion transporters (Walz, 1989;Longuemare et al., 1999). Breakdown of the Na ϩ gradient is one of the key elements in promoting cellular damage in astrocytes during energy failure (Longuemare et al., 1999). An increase in intracellular Na ϩ concentration ([Na ϩ ] i ) was found in rat spinal cord astrocytes (Rose et al., 1998), rat cortical astrocytes (Longuemare et al., 1999), and mouse cortical astrocytes (Silver et al., 1997) when these cells were exposed to glucose deprivation, NaN 3 -mediated chemical hypoxia, and simulated ischemia, respectively. Mechanisms of the rise in [Na ϩ ] i under reduced energy production conditions are not well understood. Inhibition of Na ϩ /K ϩ -ATPase activity via limited energy production results in Na ϩ accumulation (Silver et al., 1997). However, blocking voltage-gated Na ϩ channels had no effect on the rise of [Na ϩ ] i in rat spinal astrocytes during chemical hypoxia (Rose et al., 1998 reverse-mode operation of the Na ϩ /Ca 2ϩ exchanger (NCX) in neurons and causes irreversible injury during anoxia and ischemia (Li et al., 2000). An increase in intracellular Ca 2ϩ via the reverse-mode operation of NCX occurs in rat astrocytes when [Na ϩ ] i was raised by inhibition of Na ϩ /K ϩ -ATPase activity or activation of AMPA channels (Goldman et al., 1994;Smith et al., 2000). However, it is unknown whether accumulation of intracellular Na ϩ during a...

We hypothesized that high extracellular K(+) concentration ([K(+)](o))-mediated stimulation of Na(+)-K(+)-Cl(-) cotransporter isoform 1 (NKCC1) may result in a net gain of K(+) and Cl(-) and thus lead to high-[K(+)](o)-induced swelling and glutamate release. In the current study, relative cell volume changes were determined in astrocytes. Under 75 mM [K(+)](o,) astrocytes swelled by 20.2 +/- 4.9%. This high-[K(+)](o)-mediated swelling was abolished by the NKCC1 inhibitor bumetanide (10 microM, 1.0 +/- 3.1%; P < 0.05). Intracellular (36)Cl(-) accumulation was increased from a control value of 0.39 +/- 0.06 to 0.68 +/- 0.05 micromol/mg protein in response to 75 mM [K(+)](o). This increase was significantly reduced by bumetanide (P < 0.05). Basal intracellular Na(+) concentration ([Na(+)](i)) was reduced from 19.1 +/- 0.8 to 16.8 +/- 1.9 mM by bumetanide (P < 0.05). [Na(+)](i) decreased to 8.4 +/- 1.0 mM under 75 mM [K(+)](o) and was further reduced to 5.2 +/- 1.7 mM by bumetanide. In addition, the recovery rate of [Na(+)](i) on return to 5.8 mM [K(+)](o) was decreased by 40% in the presence of bumetanide (P < 0.05). Bumetanide inhibited high-[K(+)](o)-induced (14)C-labeled D-aspartate release by ~50% (P < 0.05). These results suggest that NKCC1 contributes to high-[K(+)](o)-induced astrocyte swelling and glutamate release.