Regional Cerebral Circulation And Metabolism

这是一个综合性项目，涵盖了该实验室先前开发的各种方法的开发、验证、改进和应用，以检查静止状态和功能激活时脑血流量（CBF）和能量代谢调节的基本生化和生理机制。去年完成和发表的研究表明，血管平滑肌中存在的对atp敏感的K+离子通道能够扩张血管，但对CBF的功能激活没有贡献。今年发表的另一项研究反驳了多巴胺受体参与CBF功能激活的假设。今年完成和发表的研究证实，葡萄糖利用的途径在某种程度上在星形胶质细胞和神经元之间是分开的。星形胶质细胞将葡萄糖代谢为乳酸并将其输出到神经元，神经元将乳酸氧化为CO2和H2O。然而，这些研究表明，神经元可以很容易地氧化葡萄糖和乳酸，但在动力学上更倾向于氧化来自细胞外空间的乳酸，而不是通过糖酵解在细胞内产生的丙酮酸/乳酸。星形胶质细胞也可以将葡萄糖和乳酸氧化为二氧化碳，但它们很少这样做，它们主要将葡萄糖代谢为乳酸。星形胶质细胞的能力有限是由于丙酮脱氢酶（PDH）活性有限。该酶主要以磷酸化失活形式存在于星形胶质细胞中，但它可以被二氯乙酸激活，我们发现二氯乙酸可以刺激星形胶质细胞氧化乳酸并减少其向神经元输出乳酸。在体内给药时，它对大鼠的总体行为没有产生明显的变化，并导致大脑葡萄糖利用的增加，这可能是由于葡萄糖的利用增加了，以补偿从星形胶质细胞进口乳酸的减少。这些研究的结论是，神经元和星形胶质细胞之间确实存在葡萄糖代谢的区隔化，但它既不是完全的，也不是必须的，神经元对葡萄糖和乳酸的相对氧化速率随着细胞外空间乳酸浓度的变化而变化。对两种突变小鼠的研究仍在进行中，这两种突变小鼠的α或β甲状腺受体经过基因改造，不能与l -三碘甲状腺原氨酸结合。部分研究结果已于今年公布。在β甲状腺激素受体改变的小鼠中，脑葡萄糖的利用完全正常，但在α受体功能失调的小鼠中，脑葡萄糖的利用明显地和广泛性地受到抑制，就像在出生时接受放射甲状腺切除术的动物一样。这些结果表明-甲状腺激素受体与正常的大脑发育几乎没有关系，甲状腺激素对大脑发育的影响是由-甲状腺激素受体介导的。今年发表的关于这些突变小鼠的其他研究表明，尽管大脑结构的葡萄糖利用率基线显著降低，但神经元功能激活引起的葡萄糖利用率百分比增加仍然相同，这表明基线葡萄糖利用率下降是由于突触密度降低，但其余突触功能正常。最近完成并提交发表的研究也表明，甲状腺α受体功能失调的小鼠心脏大小和心脏葡萄糖利用明显减少，而β受体突变的小鼠心脏大小略大，心脏葡萄糖利用大大增加。关于大脑中腺苷水平升高是导致自然睡眠状态的原因这一假说的研究正在进行中。有报道称，大脑腺苷水平在清醒时逐渐上升，然后在睡眠时下降到正常水平。此外，咖啡因是一种有效的非特异性腺苷通道阻滞剂，已知会干扰自然睡眠。为了在实验中提高脑腺苷水平，我们一直在给大鼠注射抑制酶的药物，这种酶能代谢通常在大脑中产生的腺苷。这两种药物分别是抑制腺苷脱氨酶的脱氧柯福霉素和抑制腺苷激酶的碘结核菌素。当静脉给药时，这些药物产生明显的动脉血压下降，这无疑是由于血管平滑肌上腺苷受体的激活引起的全身血管舒张。为了避免这种全身性低血压的并发症，我们一直在给药。到目前为止，这两种药物在脑内同时注射可以产生类似睡眠的行为状态，以及类似于快速眼动和非快速眼动睡眠状态的脑电图变化。这些研究将继续进行，以确保这些效果的可重复性，并且将在服用这些药物的大鼠中测量局部脑葡萄糖利用率，该利用率已知在非快速眼动睡眠中显着和广泛降低。

This is an omnibus project that covers a variety of studies on development, validation, refinement, and applications of methods previously developed in this laboratory to examine basic biochemical and physiological mechanisms underlying the regulation of cerebral blood flow (CBF) and energy metabolism at rest and in response to functional activation. Studies completed and published last year showed that ATP-sensitive K+ ion channels,present in vascular smooth muscle and able to dilate blood vessels, do not contibute to functional activation of CBF. Also published this year was a study that disproved the hypothesis that dopamine receptors are involved in the functional activation of CBF. Studies completed and published this year confirmed that pathways of glucose utilization are to some extent compartmentalized between astroglia and neurons. The astroglia metabolize glucose to lactate and export it to neurons, and the neurons oxidize the lactate to CO2 and H2O. These studies showed, however, that neurons can readily oxidize both glucose and lactate but have a kinetic preference to oxidize lactate derived from the extracellular space over pyruvate/lactate produced intracellularly by glycolysis. Astroglia also can oxidize both glucose and lactate to CO2, but they do so sparingly, and they metabolize glucose mainly to lactate. The limited ability of the astroglia is due to limited pyruvic dehydrogenase (PDH) activity. This enzyme exists in astroglia predominantly in the phosphorylated inactive form, but it can be activated by dichloroacetate, which we found can stimulate astroglia to oxidize lactate and diminish their export of lactate to neurons. When administered in vivo to rats, it produced no obvious changes in gross behavior and led to increases in cerebral glucose utilization, presumably due to increased utilization of glucose to compensate for the diminished import of lactate from the astroglia. The conclusion from these studies is that the compartmentalization of glucose metabolism between neurons and astroglia does exist, but it is neither complete nor obligatory, and the relative rates of neuronal oxidation of glucose and lactate vary with the lactate concentration in the extracellular space. Studies on two strains of mutant mice with either the alpha or the beta thyroid receptor genetically altered so that they could not bind L-triiodothyronine are still in progress. Part of the results were published this year. Cerebral glucose utilization was found to be completely normal in mice with the altered beta thyroid hormone receptor, but markedly and diffusely depressed in mice with the dysfunctional alpha-receptor as in animals made cretinous by radiothyroidectomy at birth. These results indicate that the beta thyroid hormone receptor has little if anything to do with normal brain development and that the effects of thyroid hormone on brain development are mediated by the alpha thyroid hormone receptor. Other studies on these mutant mice that have been published this year showed that although the baseline glucose utilization in cerebral structures is markedly reduced, the percent increases in their rates of glucose utilization evoked by neuronal functional activation reamin the same, indicating that the decreased baseline glucose utilization is due to diminished synaptic density but the remaining synapses are functionally normal. Studies recently completed and submitted for publication also showed marked reductions in heart size and cardiac glucose utilization in the mice with the dysfunctional thyroid alpha-receptors whereas heart size was slightly greater and cardiac glucose utilization were enormously increased in the mice with the mutant beta-receptor. Studies are in progress to examine the hypothesis that increased adenosine levels in brain are responsible for the induction of natural sleep states. There have been reports that brain adenosine levels progressively rise during wakefulness and then decline back to normal levels during sleep. Also, caffeine, which is a potent non-specific blocker of adenosine channels, is known to interfere with natural sleep. In order to raise brain adenosine levels experimentally, we have been infusing into rats drugs that inhibit enzymes which metabolize the adenosine normally produced in brain. These drugs are deoxycoformycin, which inhibits adenosine deaminase, and iodotubericidin, which inhibits adenosine kinase. When administered parenterally, these drugs produce marked falls in arterial blood pressure, undoubtedly due to systemic vasodilatation resulting from activation of adenosine receptors on the vascular smooth muscle. To avoid the complications of this systemic hypotension, we have been administering the drugs intracisternally. Intracisternal infusions of both drugs simultaneusly have thus far been found to produce a sleep-like behavioral state as well as EEG changes similar to those seen in REM and non-REM sleep states. These studies will be continued to assure the reproducibility of these effects, and also local cerebral glucose utilization, which is known to be markedly and diffusely reduced in non-REM sleep, will be measured in rats treated with these drugs.