Infant Rat Brain Research Articles

Formalin-Induced c-fos Expression in the Brain of Infant Rats

In the fetal, infant, and adult rat, injury induces a well-defined behavioral response and induces c-fos expression in the spinal cord dorsal horn. There is more limited information about the processing of noxious stimulation in the infant brain. We describe here the appearance of the Fos protein in the brain of fetal and infant rats following formalin-induced injury. Regions were chosen for analysis with a special focus on brain loci that express c-fos in the adult. No Fos positive cells were found in the brains of fetuses; newborns did not show increased Fos expression after formalin injection in any structure examined. At 3 and 14 days of age, there was a significant increase in Fos staining induced by formalin in the ventral lateral medulla. In contrast, paraventricular and medial dorsal nuclei of the thalamus, the paraventricular nucleus of the hypothalamus, and periaqueductal gray of the midbrain showed increased levels of Fos protein only at 14 days of age. We hypothesize that this developmental pattern is related not only to the maturation of pain perception but also to development of autonomic and defensive reactions to pain in the infant. Because the infant processes pain differently than the adult, knowledge of those differences informs pediatric clinical practice. Using Fos expression as a marker of neural activity in the rat, we show that the pattern of brain activation is immature at birth but is in place by 14 days of age.

Can't Get Next to You

The stress-response system of infant rat brains disrupts the rats' capacity for learning to maintain close proximity to mothers who are stressed themselves.

Osteopathy and (hatha) yoga

Differences and points of contact between osteopathy and yoga as regards their history and practical application are outlined. Both seek to promote healing. Yoga seeks the attainment of consciousness; osteopathy aims for providing support to health. One fundamental difference is the personal involvement of the individual in yoga. Teacher and student alike are challenged to re-examine the attitudes of mind they have adopted toward their lives. Osteopathy generally involves a relatively passive patient while the osteopath is active in providing treatment. Practical examples are used to highlight points of contact between yoga and osteopathy. The text includes a discussion of the importance of physicality and a description of ways of using it in healing processes. Furthermore, processes of attaining consciousness are outlined. Possible reductionist misconceptions in yoga and osteopathy are also pointed out. Fundamental attitudes and focus that complement each other are presented, taking the concept of stillness as a particular example.

Potential of xenon to induce or to protect against neuroapoptosis in the developing mouse brain

Drugs that suppress neuronal activity, including all general anesthetics that have been tested thus far (ketamine, midazolam, isoflurane, propofol, and a cocktail of midazolam, nitrous oxide and isoflurane), trigger neuroapoptosis in the developing rodent brain. Combinations of nitrous oxide and isoflurane, or ketamine and propofol, cause more severe neuroapoptosis than any single agent by itself, which suggests a positive correlation between increased levels of anesthesia and increased severity of neuroapoptosis. In contrast, there is evidence that the rare gas, xenon, which has anesthetic properties, protects against isoflurane-induced neuroapoptosis in the infant rat brain, while not inducing neuroapoptosis by itself. The present study was undertaken to evaluate the potential of xenon to induce neuroapoptosis or to protect against neuroapoptosis induced by isoflurane in the infant mouse brain. Seven-day-old C57BL/6 mice were exposed to one of four conditions: air (control); 0.75% isoflurane; 70% xenon; or 0.75% isoflurane + 70% xenon for four hours. For histopathological evaluation of the brains, all pups were euthanized two hours later using activated caspase-3 immunohistochemical staining to detect apoptotic neurons. Under each condition, quantitative assessment of the number of apoptotic neurons in the cerebral cortex (CC) and in the caudate/putamen (C/P) was performed by unbiased stereology. The combination of xenon + isoflurane produced a deeper level of anesthesia than either agent alone. Both xenon alone (p < 0.003 in CC; p < 0.02 in C/P) and isoflurane alone (p < 0.001 in both CC and C/P) induced a significant increase in neuroapoptosis compared to controls. The neuroapoptotic response to isoflurane was substantially more robust than the response to xenon. When xenon was administered together with isoflurane, the apoptotic response was reduced to a level lower than that for isoflurane alone (p < 0.01 in CP; marginally non-significant in CC). We conclude that xenon, in the infant mouse brain, has paradoxical properties. It triggers neuroapoptosis, and when combined with isoflurane, it increases the depth of anesthesia, and retains its own apoptogenic activity. However, it suppresses, rather than augments, isoflurane's apoptogenic activity.

PRO: Anesthesia-Induced Developmental Neuroapoptosis: Status of the Evidence

PRO: Anesthesia-Induced Developmental Neuroapoptosis: Status of the Evidence

The analysis of the shaking injury for hippocampus areas of infant rat brain by diffusion weighted imaging

The analysis of the shaking injury for hippocampus areas of infant rat brain by diffusion weighted imaging

Theanine, r-glutamylethylamide, increases neurotransmission concentrations and neurotrophin mRNA levels in the brain during lactation

Theanine (r-glutamylethylamide) is one of the major amino acid components in green tea. Recent studies suggest that theanine affects neurotransmission, especially inhibitory neurotransmission. In this study, we investigated whether theanine affects brain development in infant rats, because inhibitory neurotransmission is required for mature brain function. Mother rats were fed theanine ad libitum after confinement. The body weight gain rate of infants was not different from control infants. We detected theanine in the infant serum and measured neurotransmitter concentration and nerve growth factor (NGF) mRNA level in the infant rat brain. Some neurotransmitters, including dopamine, serotonin, glycine and GABA concentration, increased in the infant brain and NGF mRNA level increased in the cerebral cortex and hippocampus. However, these differences were lost by the end of nerve maturity. These results suggest that theanine enhanced synthesis of nerve growth factor and neurotransmitters during a nerve maturing period and promoted central nerve system maturation (CNS). Thus, theanine accelerated maturation. In conclusion, theanine may assist in healthy brain function development.

Potential of ketamine and midazolam, individually or in combination, to induce apoptotic neurodegeneration in the infant mouse brain

Recently, it was reported that anesthetizing infant rats for 6 h with a combination of anesthetic drugs (midazolam, nitrous oxide, isoflurane) caused widespread apoptotic neurodegeneration in the developing brain, followed by lifelong cognitive deficits. It has also been reported that ketamine triggers neuroapoptosis in the infant rat brain if administered repeatedly over a period of 9 h. The question arises whether less extreme exposure to anesthetic drugs can also trigger neuroapoptosis in the developing brain. To address this question we administered ketamine, midazolam or ketamine plus midazolam subcutaneously at various doses to infant mice and evaluated the rate of neuroapoptosis in various brain regions following either saline or these various drug treatments. Each drug was administered as a single one-time injection in a dose range that would be considered subanesthetic, and the brains were evaluated by unbiased stereology methods 5 h following drug treatment. Neuroapoptosis was detected by immunohistochemical staining for activated caspase-3. It was found that either ketamine or midazolam caused a dose-dependent, statistically significant increase in the rate of neuroapoptosis, and the two drugs combined caused a greater increase than either drug alone. The apoptotic nature of the neurodegenerative reaction was confirmed by electron microscopy. We conclude that relatively mild exposure to ketamine, midazolam or a combination of these drugs can trigger apoptotic neurodegeneration in the developing mouse brain.

Protection with estradiol in developmental models of apoptotic neurodegeneration

Medical measures that bear no known danger for the adult brain may trigger active neuronal death in the developing brain. Pharmacological blockade of N-methyl-D-aspartate or activation of GABA(A) receptors, blockade of voltage-dependent sodium channels, and oxygen induce widespread apoptotic neurodegeneration during the period of rapid brain growth in rodents. Because such measures are often necessary in critically ill infants and toddlers, search for adjunctive neuroprotective strategies is warranted. We report that 17beta-estradiol ameliorates neurotoxicity of drugs that block N-methyl-D-aspartate receptors, activate GABA(A) receptors, or block voltage-gated sodium channels and reduces neurotoxicity of oxygen in the infant rat brain. This neuroprotective effect is reversed by tamoxifen and cannot be reproduced by 17alpha-estradiol. 17Beta-estradiol did not affect GABA(A) or N-methyl-D-aspartate currents in hippocampal neuronal cultures, indicating that direct modulation of neurotransmitter receptor/channel properties by this compound cannot explain neuroprotective effect. 17beta-Estradiol did, however, increase levels of phosphorylated extracellular signal-regulated kinase 1/2 and AKT, suggesting that activation of these prosurvival proteins may represent one mechanism for its neuroprotective action. 17Beta-estradiol and related compounds may be neuroprotective agents suitable for use in critically ill infants and toddlers. Its supplementation may particularly help to improve neurocognitive outcome in preterm infants who are prematurely deprived of maternal estrogen.

Age-dependent effects of maternal deprivation on oxidative stress in infant rat brain

Developing brain is much more sensitive to all kind of stressors than the developed brain. Early maternal deprivation causes some behavioural and physiological effects on rats. After the birth, there is no endocrinological response to stressors between post-natal 4 and 14th days, which is called stress-hyporesponsive period (SHRP) in rats. This hypo-responsiveness is time- and stressor-specific, as some more severe stressors have been shown to induce a stress response. The present study examined the effects of maternal deprivation on oxidative stress in the hippocampus, prefrontal cortex (PFC) and striatum regions of the brain both during and after SHRP of the infant rats. The results showed that maternal deprivation in SHRP increased antioxidant enzyme activities and reduced lipid peroxidation in infant rat brain. However, by the termination of SHRP, maternal deprivation reduced enzyme activities and increased lipid peroxidation. The results indicated that infant brain might be protected in SHRP from maternal deprivation-induced oxidative stress.

The administration of cobra venom factor reduces post-ischemic cerebral injury in adult and neonatal rats

The role of complement in post-ischemic cerebral injury is incompletely understood. Therefore, experiments were designed to test the effect of complement depletion on cerebral infarct volume in adult rats and cerebral atrophy in neonatal rats. Cerebral infarcts were induced in adult rats by transient filamentous occlusion of the right middle cerebral artery (MCAO). Cerebral atrophy was induced by subjecting 7-day-old rats to ligation of the right common carotid artery followed by 2.5 h of hypoxia (8% O 2). Forty-eight hours after MCAO, coronal sections of adult brains were obtained and stained with 2,3,5-triphenyl tetrazolium chloride. The infant rat brains were removed for analysis 6 weeks after the hypoxic–ischemic insult. Volumes of infarcts and normal hemispheric parenchyma were quantified by computer-based planimetry. Twenty-four hours prior to MCAO (adults) or hypoxia–ischemia (neonates), each animal received an i.p. injection of either 1 mcg/g body weight cobra vemon factor (CVF; adult n = 11; neonatal n = 20) or normal saline (adult n = 12; neonatal n = 24). In the neonates, a second dose of CVF or saline was administered 2 days after hypoxia–ischemia. The administration of CVF significantly reduced: (1) post-ischemic cerebral infarct volume in the adults and (2) post-hypoxic–ischemic cerebral atrophy in the neonates. Therefore, complement activation augmented post-ischemic cerebral injury in adult and neonatal rats. Complement depletion induced by CVF significantly reduced post-ischemic cerebral infarct volume and atrophy in adult and neonatal rats.

Ethanol-induced neuronal apoptosis in vivo requires BAX in the developing mouse brain.

A single episode of ethanol intoxication triggers widespread apoptotic neurodegeneration in the infant rat or mouse brain. The cell death process occurs over a 6-16 h period following ethanol administration, is accompanied by a robust display of caspase-3 enzyme activation, and meets ultrastructural criteria for apoptosis. Two apoptotic pathways (intrinsic and extrinsic) have been described, either of which may culminate in the activation of caspase-3. The intrinsic pathway is regulated by Bax and Bcl-XL and involves Bax-induced mitochondrial dysfunction and release of cytochrome c as antecedent events leading to caspase-3 activation. Activation of caspase-8 is a key event preceding caspase-3 activation in the extrinsic pathway. In the present study, following ethanol administration to infant mice, we found no change in activated caspase-8, which suggests that the extrinsic pathway is not involved in ethanol-induced apoptosis. We also found that ethanol triggers robust caspase-3 activation and apoptotic neurodegeneration in C57BL/6 wildtype mice, but induces neither phenomenon in homozygous Bax-deficient mice. Therefore, it appears that ethanol-induced neuroapoptosis is an intrinsic pathway-mediated phenomenon involving Bax-induced disruption of mitochondrial membranes and cytochrome c release as early events leading to caspase-3 activation.

Heterotopic graft of infant rat brain as an ischemic model for prolonged whole-brain ischemia

By using a heterotopic brain graft model, we have made histological and electrophysiological studies of the infant rat brain after prolonged ischemia. An infant rat head which had undergone ischemia for more than 90 min, was grafted onto an adult rat by anastomosing the thoracic vessels to the femoral vessels of the host rat. Histological and histochemical studies carried out 10 days after the operation showed that the development of the hippocampus and cerebellum in the grafted brain appeared to be normal. Interneuron growth in the hippocampus and migration of the granule cells in the cerebellum had occurred to a similar extent as in control rats. Extracellular recordings in the hippocampus showed normal characteristics of the postsynaptic potentials including long-term potentiation. This heterotopic graft model would be useful for studying brain function after long periods of ischemia.

Naloxone-Precipitated Morphine Withdrawal Elicits Increases in c-fos mRNA Expression in Restricted Regions of the Infant Rat Brain

This paper is the first report of a genetic index for morphine withdrawal in infant rats. We examined the effects of naloxone (2 mg/kg) on c-fos mRNA levels in brains of infant and adult rats following repeated treatment with morphine (20 mg/kg, once daily for 5 days). One hour after a single administration of naloxone (naloxone challenge), an increase in c-fos mRNA was observed in the olfactory bulb, hypothalamus and medulla oblongata of infant rats, and in the olfactory bulb and hypothalamus, but not in the medulla oblongata of adult rats. The c-fos mRNA levels returned to control levels 6 h after the naloxone challenge. The increase in c-fos mRNA levels was followed by body weight loss in both infant and adult rats. When MK-801, a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist, was co-administered along with morphine, it inhibited the naloxone-induced increases in c-fos mRNA levels in infant rats following repeated morphine administration. These results suggest that physical dependence develops in infant rats following repeated morphine administration and that the increment of c-fos mRNA levels is a useful indicator for naloxone-precipitated morphine withdrawal in infant as well as in adult rats.

Maternal deprivation increases cell death in the infant rat brain

Prolonged separation from the mother can interfere with normal growth and development and is a significant risk factor for adult psychopathology. In rodents, separation of a pup from its mother increases the behavioral and endocrine responses to stress for the lifetime of the animal. Here we investigated whether maternal deprivation could affect brain development of infant rats via changes in the rate of cell death as measured by labeling the 3′ end of DNA fragments using terminal transferase (ApopTag). At postnatal day 12 (P12), the number of cells undergoing cell death approximately doubled in the cerebral cortex, cerebellar cortex and in several white matter tracts following 24 h of maternal deprivation. Deprivation strongly increased the number of ApopTag-labeled cells at P6 but not at P20. Stroking the infant rats only partially reversed the effects of maternal deprivation. Increased cell death in white matter tracts correlated with an induction of nerve growth factor which has been previously associated with oligodendrocyte cell death. Cell birth was either unchanged or decreased in response to deprivation. These results indicate that maternal deprivation can alter normal brain development by increasing cell death of neurons and glia, and provides a potential mechanism by which early environmental stressors may influence subsequent behavior.

Selective cholinergic denervation inhibits expression of long-term potentiation in the adult but not infant rat hippocampus

The present study assessed the role of the cholinergic systems on the expression of perforant path long-term potentiation (LTP) in rat hippocampal slices from the infant and adult brain. To denervate the cholinergic systems, 192 IgG–saporin was injected into the lateral ventricle of the infant (2-weeks-old) and adult (6-weeks-old) rat brain. There, choline acetyltransferase-immunoreactive fibers were barely detectable 2 weeks and 2 months after injection for both the groups. For the infant rats, perforant path LTP was not affected by selective cholinergic denervation; the probability of LTP development was 0.83 (five out of six slices) and 0.78 (seven out of nine slices) at 2 weeks and 2 months later in 192 IgG–saporin-treated slices, as compared with 0.83 at each period in control saline-treated slices. In contrast, the expression of the LTP was blocked by selective cholinergic denervation for the adult rats; the probability of LTP development was 0 (zero out of 10 slices) and 0.38 (three out of eight slices) at 2 weeks and 2 months later in 192 IgG–saporin-treated slices, as compared with 0.8 (eight out of 10 slices) and 0.83 (five out of six slices) at each period in control saline-treated slices. The results of the present study thus suggest that the cholinergic systems play a crucial role in the expression of LTP in the adult brain and that the denervated systems in the infant brain could be compensated by the sprouting of non-cholinergic fibers.

Nitric oxide is protective in listeric meningoencephalitis of rats.

The bacterium Listeria monocytogenes causes meningoencephalitis in humans. In rodents, listeriosis is associated with granulomatous lesions in the liver and the spleen, but not with meningoencephalitis. Here, infant rats were infected intracisternally to generate experimental listeric meningoencephalitis. Dose-dependent effects of intracisternal inoculation with L. monocytogenes on survival and activity were noted; 10(4) L. monocytogenes organisms induced a self-limiting brain infection. Bacteria invaded the basal meninges, chorioid plexus and ependyme, spread to subependymal tissue and hippocampus, and disappeared by day 7. This was paralleled by recruitment and subsequent disappearance of macrophages expressing inducible nitric oxide synthase (iNOS) and nitrotyrosine accumulation, an indication of nitric oxide (NO.) production. Treatment with the spin-trapping agent alpha-phenyl-tert-butyl nitrone (PBN) dramatically increased mortality and led to bacterial numbers in the brain 2 orders of magnitude higher than in control animals. Treatment with the selective iNOS inhibitor L-N(6)-(1-iminoethyl)-lysine (L-NIL) increased mortality to a similar extent and led to 1 order of magnitude higher bacterial counts in the brain, compared with controls. The numbers of bacteria that spread to the spleen and liver did not significantly differ among L-NIL-treated, PBN-treated, and control animals. Thus, the infant rat brain is able to mobilize powerful antilisterial mechanisms, and both reactive oxygen and NO. contribute to Listeria growth control.

Spin trapping agent, phenyl N-tert-butylnitrone, reduces nitric oxide production in the rat brain during experimental meningitis

Phenyl N-tert-butylnitrone (PBN) is a spin trapping agent previously shown to exert a neuroprotective effect in infant rat brain during bacterial meningitis. In the present study, we investigated the effect of systemic PBN administration on nitric oxide (NO) production in a rat model of experimental meningitis induced by lipopolysaccharide (LPS). We assessed the NO concentration in rat brain tissues with an electron paramagnetic resonance (EPR) NO trapping technique. In this model, rats receiving intracisternal LPS administration showed symptoms of meningitis and cerebrospinal fluid (CSF) pleocytosis. The time course study indicated that the concentration of NO in the brain reached the maximum level 8.5h after injection of LPS, and returned to the control level 24 h after the injection. When various doses of PBN (125–400 mg/kg) were injected intraperitoneally 30 min prior to LPS, NO production in the brain was reduced with increasing PBN dose (250 mg/kg suppressed 80% at 8.5h after LPS injection), and white blood cells (WBC) in CSF were significantly decreased. We concluded that reduction of NO generation during bacterial meningitis contributes to the neuroprotective effect of PBN in addition to its possible direct scavenging of reactive oxygen intermediate (ROI).

The effect of cholinergic stimulation on rat pup ultrasonic vocalizations.

As cholinergic stimulation increases vocalizations in adult rats, the present study investigated the effects of systemic oxotremorine, a cholinergic agonist, on the production of separation calls in rat pups of different ages and whether these effects are in response to central versus peripheral stimulation. The first experiment examined the dose-response effects of oxotremorine on the number of vocalizations and acoustic parameters of 10-, 15-, and 17-day-old rat pups. In contrast to other studies on adult rats, pup vocalizations were decreased while marginally changing acoustic parameters. The second experiment, using muscarinic antagonists, showed that pretreatment with atropine sulfate, which can cross the blood-brain barrier (BBB), reversed the call-reducing effect of oxotremorine whereas pretreatment with atropine methyl nitrate, which does not cross BBB, did not. Suppression of vocalizations by oxotremorine may be explained by central activation and not the peripheral effects of the drug. Dissimilar effects of cholinergic stimulation of infant and adult rat brains may be attributed to a differential role of the cholinergic system during development and maturity.

Induction of a physiologically active brain-derived neurotrophic factor in the infant rat brain by peripheral administration of 4-methylcatechol

Effects of 4-methylcatechol (4MC), a known potent stimulator of nerve growth factor (NGF) synthesis, on expression of brain-derived neurotrophic factor (BDNF) mRNA and BDNF-like immunoreactivity (BDNF-LI) was investigated in infant rat brains. A single intraperitoneal administration of 4MC caused transient increases in the levels of BDNF mRNA and BDNF-LI in neurons of the cerebral cortex from 1 to 3 h and 3 to 12 h, respectively, after the injection. Repetitive injections of 4MC to newborn rats (12-h intervals for 10 days) caused a marked and dose-dependent elevation of the level of BDNF mRNA in the whole brain besides elevating the number of cells containing calbindin D-28 and enhancing its immunoreactive intensity in the pyriform cortex and hippocampus. These findings demonstrate that 4MC stimulates de novo synthesis of BDNF in the infant rat brain, resulting in acceleration of the developmental expression of calbindin D-28.