Recombinant human erythropoietin (rEpo) is neuroprotective in neonatal models of hypoxic-ischemic brain injury. However, the optimal rEpo dose, dosing interval, and number of doses for reducing brain injury are still undetermined. We compared the neuroprotective efficacy of several subcutaneous rEpo treatment regimens. Seven-day-old rats underwent unilateral carotid ligation plus 90 min 8% hypoxia. Treatment began immediately after injury. Treatment regimens examined included 1, 3, or 7 daily subcutaneous injections of either 0 (vehicle), 2,500, 5,000, or 30,000 U/kg rEpo. Gross brain injury, neuronal apoptosis (TUNEL), and gliosis (glial fibrillary acidic protein) were assessed at 48 h or 1 wk post injury. Immunoreactive cells and brain injury were quantified for statistical comparison to vehicle controls. rEpo treatment reduced brain injury, apoptosis, and gliosis, in a dose-dependent U-shaped manner at both 48 h and 1 wk. Neither one injection of 2,500, seven injections of 5,000, or three injections of 30,000 U/kg rEpo were protective. Three doses of 5,000 and one dose of 30,000 U/kg rEpo were most protective at both time intervals. rEpo provides dose-dependent neuroprotection. Of the regimens tested, three doses of 5,000 U/kg was optimal because it provided maximal benefit with limited total exposure. (Pediatr Res 61: [451][452][453][454][455] 2007) N eonatal brain injury continues to affect the lives of thousands of infants in the United States and worldwide. In fact, perinatal asphyxia accounts for 23% of deaths globally (1). Hypothermia following hypoxia-ischemia offers promise of improved neurodevelopmental outcomes, however, thus far, the therapeutic benefit has been modest (2). When strict entry criteria are defined, up to 60% of untreated infants exposed to moderate to severe perinatal hypoxia-ischemia die or have significant neurodevelopmental sequelae (3,4). Erythropoietin (Epo), a hematopoietic cytokine known best for its erythropoietic effects, has neuroprotective effects in many animal models of brain injury, including hypoxia-ischemia, trauma, and excitotoxic injury (5-12). Although many neuroprotective strategies appeared promising in animal models, most have failed clinically (often due to toxicity). In contrast, rEpo is a safe treatment for anemia (13) and, more importantly, the neuroprotective effects of rEpo have translated well from bench to clinical trials for both amyotrophic lateral sclerosis (ALS) (14) and middle cerebral artery stroke (15).Initial studies provided no evidence that the 37 kD charged glycoprotein rEpo could cross the blood-brain barrier (BBB) (16 -18). Therefore, early rEpo neuroprotection experiments used either in vitro application (19,20) or direct intracerebral injection (21). More recently, robust neuroprotection was produced after systemic injection of high-dose rEpo (5,000 U/kg i.p.) (5). Although the neuroprotective rEpo dose is clearly above the anemia treatment range, there is no consensus as to the optimal neuroprotective dose, dose number, or in...

Recombinant human erythropoietin (rEpo) is neuroprotective in neonatal models of brain injury. Pharmacokinetic data regarding the penetration of circulating rEpo into brain tissue is needed to optimize neuroprotective strategies. We sought to determine the pharmacokinetics of rEpo given intraperitoneally or subcutaneously in plasma and brain. We hypothesized that 1) exogenous rEpo would penetrate the blood-brain barrier (BBB), 2) brain and plasma Epo would correlate, and 3) brain injury would enhance rEpo penetration. Two hundred and eighty-four 7-d-old control, sham, or brain-injured rats were treated with i.p. or s.c. rEpo (0, 250, 2500, or 5000 U/kg) and killed at scheduled intervals. Plasma and brain tissue were collected. Epo concentrations were measured by ELISA. Intraperitoneal injection yielded a faster and greater peak concentration of plasma rEpo (Tmax 3 h, Cmax 10,016 Ϯ 685 mU/mL) than s.c. injection (Tmax 9 h, Cmax 6224 Ϯ 753 mU/mL). Endogenous brain Epo was below detection even after hypoxia exposure. Systemic rEpo crossed the BBB in a dose-dependent manner, peaked in brain at 10 h, and was increased after brain injury. We conclude that highdose rEpo is detectable in brain for Ͼ20 h after a single systemic injection. These pharmacokinetic data are valuable for planning of rEpo neuroprotection experiments.

We examined whether estrogen negatively modulates cannabinoid-induced regulation of food intake, core body temperature and neurotransmission at proopiomelanocortin (POMC) synapses. Food intake was evaluated in ovariectomized female guinea pigs abdominally implanted with thermal DataLoggers and treated s.c. with the cannabinoid CB 1 /CB 2 receptor agonist WIN 55,212-2, the CB 1 receptor antagonist AM251 or their cremephor/ethanol/0.9% saline vehicle, and with estradiol benzoate (EB) or its sesame oil vehicle. Whole-cell patch clamp recordings were performed in slices through the arcuate nucleus. WIN 55,212-2 produced dose-and time-dependent increases in food intake. EB decreased food intake 8-24 h after administration, but rapidly and completely blocked the increase in consumption caused by WIN 55,212-2. EB also attenuated the WIN 55,212-2-induced decrease in core body temperature. The AM251-induced decrease in food intake was unaffected. The diminution of the WIN 55,212-2-induced increase in food intake caused by EB correlated with a marked attenuation of cannabinoid receptor-mediated decreases in glutamatergic miniature excitatory postsynaptic current frequency occurring within 10-15 minutes of steroid application. Furthermore, EB completely blocked the depolarizing shift in the inactivation curve for the A-type K + current caused by WIN 55,212-2. The EB-mediated, physiologic antagonism of these presynaptic and postsynaptic actions elicited upon cannabinoid receptor activation was observed in arcuate neurons immunopositive for phenotypic markers of POMC neurons. These data reveal that estrogens negatively modulate cannabinoid-induced changes in appetite, body temperature and POMC neuronal activity. They also impart insight into the neuroanatomical substrates and effector systems upon which these counter-regulatory factors converge in the control of energy homeostasis.

Background: During critical care in neonatal intensive care units (NICU), infants experience stressors and treatments that may produce lasting effects on adult health. An animal model simulating the NICU experience is needed to understand the impact of specific neonatal stressors. Objective: We combined approaches to develop a neonatal rat model simulating NICU stressors in order to examine the hypothesis that early stress and morphine sulfate (MS) exposure would affect development and alter adult behavior. Methods: Rat pups were exposed to stressors and given twice daily MS injections (2 mg/kg s.c.) for 5 days (postnatal days 3–7). Stress included daily maternal separation (from 08.00 to 16.00 h), hand feedings, a daily hypoxia/hyperoxia episode (100% N₂ for 8 min, then 100% O₂ for 4 min), and cold exposure (4°C for 20 min/day). Five treatment groups were formed: (1) ‘control control’ (dam reared and untreated); (2) control vehicle; (3) stress vehicle; (4) control morphine, and (5) stress morphine. Early growth and developmental indices were measured. Adult neurobehavioral tests were paw flick, passive avoidance, and forced swimming. Neonatal MS pharmacokinetics, neonatal and adult corticosterone levels, and adult hematocrit and blood pressure values were measured. Results: Neonatal stress significantly increased the mortality. Neonatal stress and MS treatment slowed early growth. Neonatal MS impaired passive avoidance learning and increased frequency, duration, and distance of forced swimming. There were no differences in corticosterone, hematocrit, or blood pressure values. Conclusions: This model simulates NICU stressors and enables measurement of acute physiological and long-term neurobehavioral indices. Neonatal MS treatment impaired the adult cognitive functioning.

HighlightsSplenosis can mimic carcinomatosis upon many imaging modalities.History of splenectomy must be considered when evaluating carcinomatosis.Scintigraphy is the preferred for confirming the presence of splenosis.