The voltage-gated calcium channel is composed of a poreforming ␣ 1 subunit and several regulatory subunits: ␣ 2 ␦, ␤, and ␥. We report here the identification of a novel ␣ 2 ␦ subunit, ␣ 2 ␦-4, from the expressed sequence tag database followed by its cloning and characterization. The novel ␣ 2 ␦-4 subunit gene contains 39 exons spanning about 130 kilobases and is colocalized with the CHCNA1C gene (␣ 1C subunit) on human chromosome 12p13.3. Alternative splicing of the ␣ 2 ␦-4 gene gives rise to four potential variants, a through d. The open reading frame of human ␣ 2 ␦-4a is composed of 3363 base pairs encoding a protein with 1120 residues and a calculated molecular mass of 126 kDa. The ␣ 2 ␦-4a subunit shares 30, 32, and 61% identity with the human calcium channel ␣ 2 ␦-1, ␣ 2 ␦-2, and ␣ 2 ␦-3 subunits, respectively. Primary sequence comparison suggests that ␣ 2 ␦-4 lacks the gabapentin binding motifs characterized for ␣ 2 ␦-1 and ␣ 2 ␦-2; this was confirmed by a [ 3 H]gabapentin-binding assay. In human embryonic kidney 293 cells, the ␣ 2 ␦-4 subunit associated with Ca V 1.2 and ␤ 3 subunits and significantly increased Ca V 1.2/␤ 3 -mediated Ca 2ϩ influx. Immunohistochemical study revealed that the ␣ 2 ␦-4 subunit has limited distribution in special cell types of the pituitary, adrenal gland, colon, and fetal liver. Whether the ␣ 2 ␦-4 subunit plays a distinct physiological role in select endocrine tissues remains to be demonstrated.

The effects of prolonged bicuculline-induced seizures on cerebral blood flow and metabolism were determined in paralyzed, mechanically ventilated neonatal dogs. Transient changes occurring early in the course of status epilepticus included significant arterial hypertension, hypocarbia, elevation of plasma norepinephrine levels, and decline in brain glucose concentration. Cerebral blood flow remained elevated throughout the 45 minutes of seizure. Determination of cerebral metabolite values by in vivo phosphorus 31 nuclear magnetic resonance spectroscopy and by in vitro enzymatic analysis of frozen brain samples showed significant decreases in the level of phosphocreatine and relatively less change in ATP values. Progressive intracellular acidosis occurred, coincident with elevation of brain lactate concentrations. We conclude that the physiological and metabolic alterations that occur during prolonged seizures are not uniform, but change with time. Any hypothesis advanced to explain the mechanism of neuronal injury during prolonged seizures must take into account these temporally related changes.

Summarytional 2 0 4 samples of arterial blood were obtained hourly forThe acute systemic and neuropathologic effects of E. coli endotoxin were determined in neonatal dogs. Administration of sublethal (LDo), moderate (LD50), or lethal (LDlo0) doses of endotoxin produced significant arterial hypotension, metabolic (lactic) acidosis, and hypoglycemia. Neuropathologic changes consisted of widespread inflammation in both grey and white matter; however, necrotic lesions were found only in forebrain white matter.determination ofplasma glucose and lactate levels by fluorometric analysis according to the method of Lowry and Passoneau (19). Body temperature was monitored by means of a rectal thermocouple (Yellow Springs) and maintained by a servo-controlled heating lamp. After insertion of the catheter, halothane was withdrawn and the animals allowed to spbntaneously breathe room air. The wound was sutured and periodically infiltrated with xylocaine, 1%. A low (LDo), moderate (LD~o), or high (LDIoo) Bacterial infections in the human infant are an important cause of subsequent mental retardation, cerebral palsy, and epilepsy (18,22,23,25). The morbidity and mortality associated with perinatal bacterial infections have been attributed to bacterial endotoxin, a lipopolysaccharide component of the bacterial cell wall. Because administration of endotoxin to laboratory animals mimics human septicemia (1,4,8,14), the systemic and neuropathologic disturbances induced by endotoxin are of considerable interest.Studies by Gilles et al. (12) have shown that administration of E. coli endotoxin to newborn kittens produces extensive brain injury. The acute systemic changes occurring during neonatal endotoxemia have not been described in detail. We, therefore, undertook a series of experiments in neonatal dogs to determine the acute neuropathologic changes occurring during endotoxemia as well as the critical physiologic and and metabolic parameters (blood pressure, heart rate, arterial blood gases, glucose, and lactate levels), which might be associated with those neuropathologic changes. Although these systemic parameters have not been studied during endotoxemia in the neonatal dog, they have been shown to be important predictors of mortality in the endotoxintreated adult dog (5,9,17,20). MATERIALS AND METHODSDose-response curve. Mongrel dogs, 1-10 days of age, were selected because their state of neurologic development at birth resembles that of the human infant of approximately 34-40 wk gestation (16). Trials were conducted in 16 animals to determine the dose of E. coli endotoxin that would result in 0, 50, and 100% mortality when injected subcutaneously. A lyophilized preparation of endotoxin from E. coli 055:B5 (Difco Laboratories) was chosen because this organism is a pathogen for human neonates, may be transmitted by nursery personnel (28), and has been previously used in neuropathologic experiments (12).Systemic responses to graded endotoxemia. The physiologic and metabolic responses of the animals were examined before and after ...

The behavioral performance of rats subjected in the neonatal period to hypoxia-ischemia at either 37 degrees C or 21 degrees C was compared to that of sham-ligated animals. Performance on complex motor tests was significantly delayed only in the hypoxic-ischemic 37 degrees C rats. However, cognitive testing disclosed significant delay of spatial learning in animals subjected to hypoxia-ischemia at 21 degrees C and those with gross infarction at 37 degrees C. There was enhanced avoidance learning in the animals with gross infarction in the hypoxia-ischemia 37 degrees C group. Hypoxic-ischemic damage in the neonatal rat at 37 degrees C results in transient delay of complex motor skills, but longer lasting cognitive changes. Hypoxia-ischemia during hypothermia produces no motor deficits, although there may be similar alterations in learning.

To investigate the relationship between neuropathologic damage and cerebral metabolic alterations during hypothermia in the neonatal animal, 7 day old Sprague-Dawley rats were subjected to unilateral common carotid artery ligation and hypoxia at 37 degrees C, 29 degrees C, and 21 degrees C. At 37 degrees C, animals had extensive infarction of tectum and ipsilateral cerebral hemisphere, and marked depletion of brain ATP. At 29 degrees C, there was no significant change in brain ATP; neuropathologic damage was limited to a few areas of necrosis in the deeper layers of cerebral cortex. No histologic injury was seen in the 21 degrees C group of rats. Profound hypothermia may prevent cerebral edema and visible neuropathologic damage associated with hypoxic-ischemic injury by decreasing cerebral metabolic demands. Moderate hypothermia confers a partial, but incomplete degree of protection; whereas during normothermia, the full extent of hypoxicischemic injury is manifest.

To clarify the effects of corticosteroids in neonatal hypoxic-ischemic brain injury, 7-day-old rats were subjected to unilateral common carotid artery ligation and hypoxia (Levine procedure) after being injected subcutaneously with saline, low-dose dexamethasone (4 mg/kg) or high-dose dexamethasone (40 mg/kg). Neither low-dose nor high-dose dexamethasone ameliorated the brain edema, lactacidemia, or hypoglycemia associated with hypoxia-ischemia. In addition, dexamethasone did not alter the pattern of neuropathologic damage or reduce the fall in brain high-energy phosphates. Finally, high-dose dexamethasone-treated animals experienced significantly more mortality than did either saline- or low-dose dexamethasone-treated animals. In this model of neonatal hypoxia-ischemia, dexamethasone did not confer any significant cerebral protection.

To explore the relationship between cerebral hemorrhage in the newborn and administration of sodium bicarbonate, we gave a standard dose of sodium bicarbonate (5 mEq/kg) to neonatal dogs and then assessed changes in cerebral blood flow, brain water content, and the blood-brain barrier. This dose of sodium bicarbonate produced no increase in blood pressure or cerebral blood flow and no alteration in blood-brain barrier. However, infusion of sodium bicarbonate did cause hyperosmolality and hypernatremia and a significant decrease in brain water content. Cerebral hemorrhage in the neonate associated with infusions of sodium bicarbonate may be related to shifts in brain water rather than to changes in blood pressure or cerebral blood flow.

No abstract