Insulin and the way the brain handles glucose.

—In living rats the concentration of insulin in the circulating blood was raised and independently of this the glucose concentration in the blood plasma was varied from hyperglycaemic to hypoglycaemic levels. Hyperglycaemia increased the influx of glucose into the brain and it also, for a limited period, increased the glucose gain by the brain. Insulin, on the other hand, did not affect influx but significantly increased the gain of glucose by the brain. It is suggested that although both hyperglycaemia and insulin can increase glucose gain by the brain they do so in entirely different ways.     

From the editors

Kuhse and Singer, the editors of this special issue of Bioethics, introduce seven articles on conflicting concepts, public policies, and standards for the determination of cardiorespiratory and brain death and the relationship of brain death to the beginning of "brain life" and to organ donation, especially from anencephalic infants. The articles are "Consciousness, the brain and what matters," by Grant Gillett; "Brain death and the anencephalic newborn," by Robert D. Truog and John C. Fletcher; "Brain death and brain life: rethinking the connection," by Jocelyn Downie; "A plea for the heart," by Martyn Evans; "The importance of knowledge and trust in the definition of death," by Bo Andreassen Rix; "Death, democracy and public ethical choice," by Reid Cushman and Soren Holm; and "Misunderstanding death on a respirator," by Tom Tomlinson.     

Mechanisms of Candida albicans trafficking to the brain

During hematogenously disseminated disease, Candida albicans infects most organs, including the brain. We discovered that a C. albicans vps51Δ/Δ mutant had significantly increased tropism for the brain in the mouse model of disseminated disease. To investigate the mechanisms of this enhanced trafficking to the brain, we studied the interactions of wild-type C. albicans and the vps51Δ/Δ mutant with brain microvascular endothelial cells in vitro. These studies revealed that C. albicans invasion of brain endothelial cells is mediated by the fungal invasins, Als3 and Ssa1. Als3 binds to the gp96 heat shock protein, which is expressed on the surface of brain endothelial cells, but not human umbilical vein endothelial cells, whereas Ssa1 binds to a brain endothelial cell receptor other than gp96. The vps51Δ/Δ mutant has increased surface expression of Als3, which is a major cause of the increased capacity of this mutant to both invade brain endothelial cells in vitro and traffic to the brain in mice. Therefore, during disseminated disease, C. albicans traffics to and infects the brain by binding to gp96, a unique receptor that is expressed specifically on the surface of brain endothelial cells.     

Utilization of High-Density Lipoprotein Sphingomyelin by the Developing and Mature Brain in the Rat

Utilization of very long chain saturated fatty acids by brain was studied by injecting 20-day-old and adult rats with high-density lipoprotein containing [stearic or lignoceric acid-14C, (methyl-3H)choline]sphingomyelin. Labeling was followed for 24 h. Very small amounts of 14C were recovered in the brain of all rats, and there was no preferential uptake of lignoceric acid. Approximately 20% of the entrapped 14C was located in the form of unchanged sphingomyelin 24 h after injection. This result shows that the rat brain utilizes very little very long chain fatty acids (greater than or equal to 20 C atoms) from high-density lipoprotein sphingomyelin, even during the myelinating period. The [3H]choline moiety from sphingomyelin was recovered in brain phosphatidylcholine in a higher proportion in comparison with the 14C uptake. The brain 3H increased throughout the studied period in all experiments, but was much higher in the myelinating brain than in the mature brain. From the radioactivity distribution in liver and plasma lipids, it is clear that the choline 3H in the brain originates from either double-labeled phosphatidylcholine of lipoproteins or tritiated lysophosphatidylcholine bound to albumin, both synthesized by the liver.     

The correlation of changes of the optic nerve diameter in the acute retrobulbar neuritis with the brain changes in multiple sclerosis

The aim of this paper is to compare diameter of healthy and affected optic nerve determined by ultrasound with brain lesions in acute retrobulbar neuritis in patients with multiple sclerosis. In this prospective study 20 patients with multiple sclerosis and acute retrobulbar neuritis were examined. Optic nerve diameter was measured by ultrasound. Brain lesions were detected by magnetic resonance. Correlation between demyelinating lesions of the brain in multiple sclerosis and optic nerve diameter was tested by Kruskal-Wallis test. Significant difference in diameter between healthy and affected optic nerve in acute retrobulbar neuritis was found. Demyelinating brain changes examined by magnetic resonance revealed periventricular lesions, subcortical lesions and lesions in corpus callosum. There is statistically significant correlation between optic nerve diameter and number of brain lesions in multiple sclerosis, p < 0.05. Diameter of optic nerve in retrobulbar neuritis measured by ultrasound correlates with brain lesions detected by magnetic resonance in multiple sclerosis.     

Hedgehog-Gli signalling and the growth of the brain

The development of the vertebrate brain involves the creation of many cell types in precise locations and at precise times, followed by the formation of functional connections. To generate its cells in the correct numbers, the brain has to produce many precursors during a limited period. How this is achieved remains unclear, although several cytokines have been implicated in the proliferation of neural precursors. Understanding this process will provide profound insights, not only into the formation of the mammalian brain during ontogeny, but also into brain evolution. Here we review the role of the Sonic hedgehog-Gli pathway in brain development. Specifically, we discuss the role of this pathway in the cerebellar and cerebral cortices, and address the implications of these findings for morphological plasticity. We also highlight future directions of research that could help to clarify the mechanisms and consequences of Sonic hedgehog signalling in the brain.     

Social Cognition,the Male Brain and the Autism Spectrum

Behavioral studies have shown that, at a population level, women perform better on tests of social cognition and empathy than men. Furthermore Autism Spectrum Disorders (ASDs), which are characterized by impairments in social functioning and empathy, occur more commonly in males than females. These findings have led to the hypothesis that differences in the functioning of the social brain between males and females contribute to the greater vulnerability of males to ASD and the suggestion that ASD may represent an extreme form of the male brain. Here we sought to investigate this hypothesis by determining: (i) whether males and females differ in social brain function, and (ii) whether any sex differences in social brain function are exaggerated in individuals with ASD. Using fMRI we show that males and females differ markedly in social brain function when making social decisions from faces (compared to simple sex judgements) especially when making decisions of an affective nature, with the greatest sex differences in social brain activation being in the inferior frontal cortex (IFC). We also demonstrate that this difference is exaggerated in individuals with ASD, who show an extreme male pattern of IFC function. These results show that males and females differ significantly in social brain function and support the view that sex differences in the social brain contribute to the greater vulnerability of males to ASDs.     

Changes in the nitric oxide level in the rat liver as a response to brain injury

Liver disturbances stimulate inflammatory reaction in the brain but little is known if injury to the brain can significantly influence liver metabolism. This problem is crucial in modern transplantology, as the condition of the donor brain seems to strongly affect the quality (viability) of the graft, which is often obtained from brain-dead donors, usually after traumatic brain injury. Because nitric oxide is one of the significant molecules in brain and liver biology, we examined if brain injury can affect NO level in the liver. Liver samples of Wistar rats were collected and studied with EPR NO-metry to detect NO level changes at different time points after brain injury. Shortly after the trauma, NO level in the liver was similar to the control. However, later there was a significant increase in the NO content in the livers starting from the 2nd day after brain injury and lasting up to the 7th day. It seems that the response to a mechanical brain injury is of the systemic, rather than local character. Therefore brain metabolism disturbances can influence liver metabolism at least by stimulating the organ to produce NO.     

Superoxide radical formation and associated biochemical alterations in the plasma membrane of brain, heart, and liver during the lifetime of the rat.

Plasma membrane samples from rat brain, heart, and liver were examined for biochemical changes with age. A rise in superoxide radical (SOR) levels was followed by increases in thiobarbituric acid reactive substances and decreases in membrane fluidity with age. The earliest rise in SOR formation appeared in the plasma membrane from the brain. With age, protein synthesis also decreased significantly in tissue homogenates from brain and heart but was unchanged in the liver. Exposure of plasma membrane samples to in vitro-elevated SOR levels stimulated formation of lipid peroxides, as indicated by the thiobarbituric acid test, and resulted in a decrease in membrane fluidity in each tissue and in a decline in protein synthesis in brain and heart. Changes in brain lipid peroxidation and in membrane fluidity in brain and heart as a result of SOR supplementation were further enhanced due to age. In addition, the mechanism of SOR formation was examined in plasma membrane samples from the brain. SOR generation was Ca(2+)-sensitive, blocked by superoxide dismutase or vitamin E and inhibited by both indomethacin, a cyclooxygenase inhibitor, and bromophenacyl bromide, a phospholipase A2 inhibitor. These results show significant increases in SOR formation and biochemical alterations in plasma membranes from brain, heart, and liver in aging rats. SOR formation appears to be enzyme-mediated and elevated levels of this oxygen radical could be involved in membrane breakdown in older rats.     

Unique Features of the Insulin Receptor in Rat Brain

We examined the structure of the affinity-labeled insulin receptors in rat brain, rat liver, and human IM-9 lymphocytes using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In gels run under reducing conditions, the alpha-subunit of the insulin receptor in brain had an apparent Mr of 127,000 distinctly lower than that seen in both rat liver and human lymphocytes (apparent Mr = 136,000). Exposure to neuraminidase increased the electrophoretic mobility of the liver receptor, but had no effect on the insulin receptor in brain. The carbohydrate moieties of the insulin receptors in rat brain and liver were further examined by chromatography on wheat-germ agglutinin agarose. The receptors in both tissues adsorbed to the wheat-germ agglutinin; elution with 0.3 M N-acetyl glucosamine resulted in slightly better recovery of the brain than of the liver receptor. Exposure to neuraminidase virtually abolished the interaction of the liver receptor with the lectin, whereas adsorption of the brain receptor was unaffected by neuraminidase. These results indicate that the insulin receptor in brain is distinguished from those in peripheral tissues by structural alterations, including changes in the carbohydrate moiety of the receptor. Such alterations contrast sharply with the previously observed similarities in insulin binding properties between insulin receptors in brain and other tissues. The implications of such structural alterations for the program of insulin action expressed by the receptors in brain remain to be explored.     

The distribution of aluminum into and out of the brain

The extent, rate and possible mechanism(s) by which aluminum enters and is removed from the brain are presented. Introduction of Al into systemic circulation as Al.transferrin, the predominant Al species in plasma, resulted in about 7 x 10(-5) of the dose in the brain 1 day after injection. This brain Al entry could be mediated by transferrin-receptor-mediated endocytosis (TfR-ME). When Al.citrate, the predominant small molecular weight Al species in blood plasma, is introduced systemically, Al rapidly enters the brain. The rate of Al.citrate brain influx suggests a more rapid process than mediated by diffusion or TfR-ME. The question has been raised: "Is the brain a 'one-way sink' for aluminum?". Clinical observations are a basis for this suggestion. Rat brain 26Al concentrations decreased only slightly from 1 to 35 days after systemic 26Al injection, in the absence or presence of the aluminum chelator desferrioxamine, suggesting prolonged brain Al retention. However, studies of brain and blood extracellular Al at steady state, using microdialysis, suggest brain Al efflux exceeds influx, suggesting carrier-mediated brain Al efflux. The predominant brain extracellular fluid Al species is probably Al.citrate. The hypothesis that brain Al efflux, presumably of Al.citrate, is mediated by the monocarboxylate transporter was tested and supported. Although some Al that enters the brain is rapidly effluxed, it is suggested that a fraction enters brain compartments within 24 h from which it is only very slowly eliminated.     

Relationships between the brain and the immune system

The concept that the brain can modulate activity the immune system stems from the theory of stress. Recent advances in the study of the inter-relationships between the central nervous system and the immune system have demonstrated a vast network of communication pathways between the two systems. Lymphoid organs are innervated by branches of the autonomic nervous system. Accessory immune cells and lymphocytes have membrane receptors for most neurotransmitters and neuropeptides. These receptors are functional, and their activation leads to changes in immune functions, including cell proliferation, chimiotactism and specific immune responses. Brain lesions and stressors can induce a number of changes in the functioning of the immune system. All these changes are not necessarily mediated by the neuroendocrine system. They can also be dependent on autonomic nerve function. The communication pathways that link the brain to the immune system are normally activated by signals from the immune system, and they serve to regulate immune responses. These signals originate from accessory immune cells such as monocytes and macrophages and they are represented mainly by proinflammatory cytokines. Proinflammatory cytokines produced at the periphery act on the brain via two major pathways: (1) a humoral pathway allowing pathogen specific molecular patterns to act on Toll-like receptors in those brain areas that are devoid of a functional blood-brain barrier, the so-called circumventricular areas; (2) a neural pathway, represented by the afferent nerves that innervate the bodily site of infection and injury. In both cases, peripherally produced cytokines induce the expression of brain cytokines that are produced by resident macrophages and microglial cells. These locally produced cytokines diffuse throughout the brain parenchyma to act on target brain areas so as to organise the central components of the host response to infection (fever, neuroendocrine activation, and sickness behavior).     

Properties allowing the attribution to gamma-hydroxybutyrate the quality of neurotransmitter in the central nervous system

gamma-Hydroxybutyrate (GHB) fulfills the main criteria of a neurotransmitter: it is unevenly distributed in C.N.S.; it is synthesized from succinic semi-aldehyde by a specific semi-aldehyde succinic reductase localized in neurons, in some dendrites and synaptic terminals; GHB is released by tissue slice depolarization, this release being reduced by 50-60% in a Ca++ free medium. Tetrodotoxin and verapamil strongly inhibited the depolarization evoked-release; high affinity heterogenously distributed binding sites for gamma-hydroxybutyrate exist in the brain. This binding does not require Na+. The bound gamma-hydroxybutyric acid is not displaceable by GABA or GABA agonists. Binding sites are enriched in the synaptosomal fraction; after micro-iontophoretic application, GHB exerts a depressant action on nigral and neocortical cells which is resistant to the presence of bicuculline methiodide. In neuronal cultures, GHB causes a hyperpolarization similar to that produced by GABA; high affinity uptake system for GHB exists both in purified plasma membrane vesicles and in brain tissue slices. This uptake is dependent on an Na+ gradient and is inhibited by ouaba?n and dinitrophenol; GABA does not modify GHB uptake by rat brain slices; GABA derived GHB has a turnover time almost three times faster than that of whole brain serotonin, 6-8 times as rapid as that of whole brain dopamine and 13-19 times as rapid as that of whole brain norepinephrine.     

川金丝猴脑的动脉供应

为了探讨川金丝猴脑动脉供应的形态学特征,为脑生物学研究提供结构基础,用血管铸型和组织透明方法追踪观察了川金丝猴幼体脑动脉的来源和分支分布。结果表明川金丝猴与人脑的动脉供应基本相同,也由颈内动脉和椎动脉供应。上述动脉的分支于垂体周围形成大脑动脉环。颈内动脉通过大脑前动脉和大脑中动脉主要供应大脑半球前部的血液,椎动脉参与形成基底动脉、小脑动脉系和大脑后动脉,供应脑干、小脑和大脑后部的血液。另外,川金丝猴幼体左、右大脑前动脉间缺少前交通动脉。     

脑水肿时紧密连接蛋白occludin及AQP4的表达变化

目的:采用枕大池内注入脂多糖(lipopolysaccharides,LPS)的方法建立大鼠脑水肿模型,观察脑组织病理形态学变化,脑组织含水量(brain water content,BWC),血脑屏障(blood brain barrier,BBB)的紧密连接蛋白Occludin和水通道蛋白-4(aquaporin 4,AQP4)表达水平的动态变化,研究AQP4及Occludin与脑水肿形成的关系,及其可能的作用机制,为临床脑水肿的治疗提供理论依据。方法:选用Wistar健康成年大鼠,随机分为正常对照组,生理盐水组和脂多糖组,后两组的观察时间点选定于造模后3 h、6h、12 h、24 h、72 h。采用经皮穿刺枕大池内注入脂多糖的方法制备脑水肿动物模型,正常对照组、生理盐水组及脂多糖组分别于各时间点进行开颅取脑,测定脑组织含水量,通过HE染色法观察脑组织的病理形态学变化,应用Western blot方法检测occludin的表达变化。应用RT-PCR技术测定脑组织内AQP4mRNA的表达变化。结果:生理盐水组各时间点中有少量AQP4mRNA及occludin蛋白的表达,与正常对照组之间无显著性差异;脂多糖组在造模后3 hAQP4的mRNA表达开始增加,6-12 h达高峰,此后明显下降,随后表达开始减弱,24-72 h表达显著低于生理盐水组;occludin蛋白表达下降出现于造模后3 h,12-24 h下降更明显,72 h表达开始升高。结论:枕大池内注入脂多糖(LPS)所建立脑水肿模型中,脑组织含水量及血脑屏障通透性增加,病理学特点是血管源性脑水肿出现早且持久,后期伴有细胞毒性脑水肿的改变。AQP4早期表达增强是胶质细胞的适应性反应,与血脑屏障的破坏有关,促进了血管源性脑水肿的发生。后期AQP4表达减弱是机体内在防御机制的表现,同时又促进细胞毒性脑水肿的形成。occludin在脑组织中表达量随脑水肿的加重而降低,即与脑水肿的程度呈负相关,目前认为这与脑水肿时内皮细胞通透性增加,血脑屏障的通透性改变,导致occludin的表达下调有关,促进了血管源性脑水肿的发生。针对以上特点,我们可以进一步研究调控AQP4及occludin表达的药物,从而减轻脑损伤后脑水肿的程度,为脑水肿的治疗提供新的临床策略。     

Cytochrome P450s of the 4A Subfamily in the Brain

Abstract: Members of the P450 4A subfamily are key enzymes in the synthesis and degradation of metabolites of arachidonic acid, which are of physiological importance in the brain. In the rat, four members of this subfamily, 4A1, 4A2, 4A3, and 4A8, have been described. In this study, the expression of members of the 4A subfamily in the rat brain has been examined by PCR amplification, by western and northern blotting, and by protein N-terminal sequencing. With PCR all four members of the subfamily were detectable in the liver and kidney. P450 4A1 was found exclusively in the liver and kidney, whereas P450 4A2 was detectable in all the tissues tested, including the lung, seminal vesicles, prostate, cerebral cortex, hypothalamic preoptic area, cerebellum, and brainstem. The tissue distribution of P450 4A3 was similar to that of 4A2 except that it was not detectable in seminal vesicles. A P450 4A8-specific fragment was amplified from the kidney, liver, and prostate and weakly from the cerebral cortex but not from other brain regions. Despite the evidence of their presence by PCR, no members of the 4A family were detectable on northern blots with mRNA from the brain. On western blots a P450 4A-specific antiserum recognized a band in P450 fractions prepared from the brain. The intensity of the signal with 30 pmol of P450 from the brain was similar to that with 10 pmol of liver microsomal P450. The brain P450 was extracted from 1 g of brain, whereas the 10 pmol of liver P450 is the equivalent of 1 mg of liver. This suggests a brain content of 4A P450 that is 0.1% of that in the liver. N-terminal sequencing of the protein bands in the brain P450 fraction revealed the presence of both P450 4A8 and 4A3. These data show the presence in the brain of forms of P450 whose level of mRNA is too low to be detected on northern blots. The specificity of tissue distribution shows that this is not just a nonspecific background level of expression and suggests a role of brain P450 in the synthesis and degradation of arachidonic acid metabolites.     

Transport of thyroxine across the blood-brain barrier is directed primarily from brain to blood in the mouse

The role of the blood-brain barrier (BBB) in the transport of thyroxine was examined in mice. Radioiodinated ("hot") thyroxine (hT4) administered icv had a half-time disappearance from the brain of 30 min. This increased to 60 min (p less than 0.001) when administered with 211 pmole/mouse of unlabeled ("cold") thyroxine (cT4). The Km for this inhibition of hT4 transport out of the brain by cT4 was 9.66 pmole/brain. Unlabeled 3,3',5 triiodothyronine (cT3) was unable to inhibit transport of hT4 out of the brain, although both cT3 (p less than 0.05) and cT4 (p less than 0.05) did inhibit transport of radioiodinated 3,3',5 triiodothyronine (hT3) to a small degree. Entry of hT4 into the brain after peripheral administration was negligible and was not affected by either cT4 nor cT3. By contrast, the entry of hT3 into the brain after peripheral administration was inhibited by cT3 (p less than 0.001) and was increased by cT4 (p less than 0.01). The levels of the unlabeled thyroid hormones administered centrally in these studies did not affect bulk flow, as assessed by labeled red blood cells (99mTc-RBC), or the carrier-mediated transport of iodide out of the brain. Likewise, the vascular space of the brain and body, as assessed by 99mTc-RBC, was unchanged by the levels of peripherally administered unlabeled thyroid hormones. Therefore, the results of these studies are not due to generalized effects of thyroid hormones on BBB transport. The results indicate that in the mouse the major carrier-mediated system for thyroxine in the BBB transports thyroxine out of the brain, while the major system for triiodothyronine transports hormone into the brain.     

Rapid transferrin efflux from brain to blood across the blood-brain barrier

The brain efflux index method is used to examine the extent to which transferrin effluxes from brain to blood across the blood-brain barrier (BBB) following intracerebral injection. Whereas high-molecular-weight dextran is nearly 100% retained in brain for up to 90 min after intracerebral injection in the Par2 region of the parietal cortex of brain, there is rapid efflux of transferrin from brain to blood across the BBB. The efflux of apotransferrin is 3.5-fold faster than the efflux of holo-transferrin. The brain to blood efflux of apotransferrin is completely saturable by unlabeled transferrin, but is not inhibited by other plasma proteins. These studies provide evidence for reverse transcytosis of transferrin from brain to blood across the BBB. As circulating transferrin is known to undergo transcytosis across the BBB in the blood-to-brain direction, these studies support the model of bidirectional transcytosis of transferrin through the BBB in vivo.     

The Nature and Composition of the Internal Environment of the Developing Brain

1. The fetal brain develops within its own environment, which is protected from free exchange of most molecules among its extracellular fluid, blood plasma, and cerebrospinal fluid (CSF) by a set of mechanisms described collectively as brain barriers.2. There are high concentrations of proteins in fetal CSF, which are due not to immaturity of the blood–CSF barrier (tight junctions between the epithelial cells of the choroid plexus), but to a specialized transcellular mechanism that specifically transfers some proteins across choroid plexus epithelial cells in the immature brain.3. The proteins in CSF are excluded from the extracellular fluid of the immature brain by the presence of barriers at the CSF–brain interfaces on the inner and outer surfaces of the immature brain. These barriers are not present in the adult.4. Some plasma proteins are present within the cells of the developing brain. Their presence may be explained by a combination of specific uptake from the CSF and synthesis in situ. 5. Information about the composition of the CSF (electrolytes as well as proteins) in the developing brain is of importance for the culture conditions used for experiments with fetal brain tissue in vitro, as neurons in the developing brain are exposed to relatively high concentrations of proteins only when they have cell surface membrane contact with CSF.6. The developmental importance of high protein concentrations in CSF of the immature brain is not understood but may be involved in providing the physical force (colloid osmotic pressure) for expansion of the cerebral ventricles during brain development, as well as possibly having nutritive and specific cell development functions.     

Salivary lactoferrin is transferred into the brain via the sublingual route

Lactoferrin (LF) is produced by exocrine glands including salivary gland, and has various functions including infection defense. However, the transfer of LF from peripheral organs into the brain remains unclear. To clarify the kinetics of salivary LF (sLF), we investigated the consequences of sialoadenectomy and bovine LF (bLF) sublingual administration in rats. The salivary glands were removed from male Wistar rats, and we measured rat LF levels in the blood and brain at 1 week post-surgery. We also examined the transfer of LF into the organs of the rats after sublingual administration of bLF. Rat LF levels in the blood and brain were significantly reduced by sialoadenectomy. Sublingual bLF administration significantly increased bLF levels in the brain, which then decreased over time. These results indicate that LF is transferred from the sublingual mucosa to the brain, in which favorable effects of sLF on brain will be expected via the sublingual mucosa.