羟丁酸钠对缺血缺氧性脑损伤新生大鼠大脑皮层神经细胞凋亡的影响

目的 观察羟丁酸钠对缺血缺氧性脑损伤新生大鼠大脑皮层神经细胞凋亡的影响。方法 新生7dSD大鼠随机分为假手术组（S组）、生理盐水对照组（C组）和羟丁酸钠组（71、坦和档组）。每组按Ⅲ（缺血缺氧）后不同时间点进一步分为1h、3h、1d、3d、7d5个亚组（n=6）。按Rice法制作缺血缺氧性脑损伤模型。C、γ1、γ2和γ3组HI后即腹腔分别注射生理盐水0．2ml／10g、羟丁酸钠50、100、200mg／kg，3次／日；S组术后吸空气2h，腹腔注射生理盐水0．2ml／10g，3次／日。TUNEL染色法检测HI后各时间点大脑皮层凋亡神经细胞。结果 HI后1h～7d，C、γ1、γ2、γ3组凋亡神经细胞数高于S组（P〈0．05），且于HI后1d达高峰；HI后3h-3d，C、γ3组凋亡神经细胞数多于γ1、γ2组（P〈0．05），而C组与γ3组之间凋亡神经细胞数差异无统计学意义（P〉0．05）；HI后7d，γ1组凋亡神经细胞数多于让组（P〈0．05）。结论 50、100mg／kg羟丁酸钠能抑制缺血缺氧性脑损伤新生大鼠大脑皮层神经细胞凋亡，且100mg／kg效果较好。     

Intensity-dependent regional cerebral blood flow during 1-Hz repetitive transcranial magnetic stimulation (rTMS) in healthy volunteers studied with H215O positron emission tomography: I. Effects of primary motor cortex rTMS.

BACKGROUND: Repetitive transcranial magnetic stimulation (rTMS) affects the excitability of the motor cortex and is thought to influence activity in other brain areas as well. We combined the administration of varying intensities of 1-Hz rTMS of the motor cortex with simultaneous positron emission tomography (PET) to delineate local and distant effects on brain activity. METHODS: Ten healthy subjects received 1-Hz rTMS to the optimal position over motor cortex (M1) for producing a twitch in the right hand at 80, 90, 100, 110, and 120% of the twitch threshold, while regional cerebral blood flow (rCBF) was measured using H(2)(15)O and PET. Repetitive transcranial magnetic stimulation (rTMS) was delivered in 75-pulse trains at each intensity every 10 min through a figure-eight coil. The regional relationship of stimulation intensity to normalized rCBF was assessed statistically. RESULTS: Intensity-dependent rCBF increases were produced under the M1 stimulation site in ipsilateral primary auditory cortex, contralateral cerebellum, and bilateral putamen, insula, and red nucleus. Intensity-dependent reductions in rCBF occurred in contralateral frontal and parietal cortices and bilateral anterior cingulate gyrus and occipital cortex. CONCLUSIONS: This study demonstrates that 1-Hz rTMS delivered to the primary motor cortex (M1) produces intensity-dependent increases in brain activity locally and has associated effects in distant sites with known connections to M1.     

Single unit activity in the auditory cortex of a monkey performing a short term memory task

Summary Short term memory to tones (STMT) was investigated by recording single unit activity in the auditory cortex of a behaving monkey. The activity of each unit was studied in two behavioral conditions: a) During task performance, the monkey had to compare two tones separated by one second of silence (inter-stimulus interval), b) During a nonperforming period; the monkey heard the two tones but did not respond behaviorally. It was noted that the firing rate of many units during the inter-stimulus interval (ISI) was dependent on the frequency of the first tone. Such dependency was observed even towards the end of the ISI, both during task performance trials (50% of the units) and during the nonperforming period (32% of the units). The activity of these units could be the basis of STMT in both of these behavioral states. In 65% of all the units tested, the responses during the ISI were of a higher magnitude in the performance period than were the responses in the non-performance period. The activity of these units may be related either to general processes such as attention and expectation or to short-term memory processes. During task performance, the responses of 23% of the units to the second tone were dependent on whether its frequency was identical to that of the first tone. Such dependency was never observed during the non-performing period. These units may detect similarity or non similarity between two tones presented one second apart. Periodic patterns of firing were not found in the study, thus suggesting that the ISI responses were not generated by reverberatory activity in simple closed loops. On the basis of these results, several alternative mechanisms of STMT are suggested.     

Variation in the ability of neuroleptics to block the inhibitory influence of dopaminergic neurons on the activity of cells in the rat prefrontal cortex

The stimulation of the ventro-medial mesencephalic tegmentum (VMT) induces an inhibition of the spontaneous activity of prefrontal cortical cells and blocks the excitatory responses evoked by the stimulation of the medio-dorsal nucleus of the thalamus (MD). This effect is mediated by the activation of the mesocortical dopaminergic (DA) system. In the present study, the influence of the systemic administration of several neuroleptics on the inhibition of prefrontal cortical cells induced by VMT stimulation has been analyzed in ketamine anaesthetised rats. The acute IP administration of fluphenazine (2 mg/kg), spiroperidol (2 mg/kg) or (+/-)sulpiride (100 mg/kg) reversed the inhibitory responses. Moreover, the number of cortical cells inhibited by VMT stimulation was considerably decreased after these treatments. Surprisingly, neither haloperidol at any dose used (0.1 to 0.5 mg/kg IV or 0.5 to 5 mg/kg IP) nor levomepromazine (25 mg/kg IP) nor the long acting neuroleptic, pipotiazine palmitic ester (32 mg/kg SC) blocked the inhibitory effect of VMT stimulation and in fact they lengthened the duration of the inhibition. Finally, the inhibition of MD evoked spikes in prefrontal cortical cells produced by VMT stimulation was no longer observed after sulpiride but persisted after haloperidol administration. Our findings confirm that the mesocortico-prefrontal DA neurons exert an inhibitory influence on target cells but they reveal differences in the efficacy of neuroleptics in blocking this effect.     

Pharmacological characterization of the receptor mediating the adrenergic inhibition of responses to substance P in the cingulate cortex

The excitatory responses of neurones in the anterior cingulate cortex of the rat to iontophoretically applied substance P (SP) are reduced by noradrenaline (NA) applied iontophoretically or released from noradrenergic pathways. In order to determine the receptor involved in this inhibitory effect we have studied the effects of a number of receptor-specific adrenergic agonists and antagonists on responses of cingulate neurones to SP in rats anaesthetized with chloral hydrate. Low iontophoretic currents (0-15 nA) of NA, adrenaline and the beta-agonist, clenbuterol, all strongly reduced responses to SP. Isoprenaline was also effective but less consistently so, although problems were experienced with its iontophoretic release from micropipettes. The alpha 1-agonists, phenylephrine and methoxamine were also able to reduce responses to SP. However, this reduction required higher iontophoretic currents (15-60 nA) and was associated with depressant effects on baseline firing rate. The alpha 2-agonist clonidine was only weakly active at high currents and this too was associated with depression of baseline firing. Similar weak effects were noted with dopamine. The inhibitory effects of NA on SP responses were convincingly blocked or reversed by the beta-antagonist, practolol, but not by the alpha 1-antagonist, prazosin. The reduction of SP responses by phenylephrine was also blocked by practolol but unaffected by prazosin. Finally, reduction of SP excitations by activation of the coeruleocortical pathway was also blocked by practolol applied iontophoretically to the cortical cells. These results are consistent with the hypothesis that the effect of NA on SP responsiveness in the cingulate cortex is mediated by beta-adrenoreceptors.     

Visual recognition impairment follows ventromedial but not dorsolateral prefrontal lesions in monkeys

Visual recognition in monkeys appears to involve the participation of two limbothalamic pathways, one including the amygdala and the magnocellular portion of the medial dorsal nucleus (MDmc) and the other, the hippocampus and the anterior nuclei of the thalamus (Ant N). Both MDmc and Ant N project, in turn, to the prefrontal cortex, mainly to its ventral and medial portions. To test whether the prefrontal projection targets of the two limbothalamic pathways also participate in memory functions, performance on a variety of learning and memory tasks was assessed in monkeys with lesions of the ventromedial prefrontal cortex (Group VM). Normal monkeys and monkeys with lesions of dorsolateral prefrontal cortex (Group DL) served as controls. Group VM was severely impaired on a test of object recognition, whereas Group DL did not differ appreciably from normal animals. Conversely, the animals in Group VM were able to learn a spatial delayed response task, whereas 2 of the 3 animals in Group DL could not. Neither group was impaired in the acquisition of visual discrimination habits, even though the successive trials on a given discrimination were separated by 24-h intervals. The patterns of deficit suggest that ventromedial prefrontal cortex constitutes another station in the limbothalamic system underlying cognitive memory processes, whereas the dorsolateral prefrontal cortex lies outside this system. The results support the view that the classical delayed-response deficit observed after dorsolateral prefrontal lesions represents a perceptuo-mnemonic impairment in spatial functions selectively rather than a memory loss of a more general nature.     

Modification of the Na,K-pump of glial cells within cobalt-induced epileptogenic cortex of rat

The characteristics of a glial Na⁺,K⁺-pump dependent on extracellular K⁺ within epileptogenic cortex were studied electrophysiologically, biochemically and histochemically in vitro using slices from cobalt-induced epileptogenic cortex of rat. When the extracellular K⁺ concentration ([K⁺]_o) was varied between 4 and 40 mM, the mean slope of membrane potential plotted against [K⁺]_o was about 57 mV in glia from the normal cortex (tissue A) and about 44 mV in glia from the epileptogenic cortex (tissue B); whereas no significant difference in the resting membrane potential of these tissues was observed. In glia from tissue B, a marked transient hyperpolarization above control level was caused by replacement of elevated [K⁺]_o with the normal medium. Ouabain abolished these phenomena observed in glia from tissue B, but had no effect on the membrane potential during normal [K⁺]_o. Reduction of extracellular Na⁺, Ca²⁺ and Cl⁻ did not significantly affect the membrane potential of glia from either tissue. In tissue A, the cells marked by intracellular injection of horseradish peroxidase after intracellular recording were protoplasmic astrocytes; in tissue B, fibrous astrocytes with abnormal processes predominated. K⁺-dependent stimulation of Na⁺,K⁺-ATPase activity of the astrocyte-enriched fraction and its membrane preparation from tissue B was much larger than that from tissue A. A certain amount of the reaction product of K⁺-pNPPase activity was seen on glial plasma membrane within tissue B but not on that from tissue A. The above findings suggest that a glial Na⁺,K⁺-pump within actively firing epileptogenic cortex may be modified to increase in its activity.     

Characterization of ramified microglia in tissue culture: pinocytosis and motility

Functional properties of ramified microglia were investigated in primary cultures of rat cerebral cortical cells. These microglia could be readily identified in both fixed and living cultures through previously established features. Based on their destruction by 5 mM L-leucine methyl ester, a high level of intrinsic endocytotic activity was established. When cultures were incubated with fluorescent latex beads to assess phagocytosis, little or no such activity was exhibited by ramified cells. However, when cultures were incubated with dyes or other soluble tracer compounds, these cells always exhibited labeling. This labeling was selective for ramified microglia in the cultures and was demonstrated using a variety of compounds, including trypan blue, lucifer yellow, horseradish peroxidase (HRP), and India ink. Intracellular label could be observed in vesicular structures; this localization corresponded to an active cellular process. Also, cellular labeling was inhibited by the presence of colchicine. These features supported the inference that the labeling was attributable to pinocytosis, and this process appeared to account for the vast majority of endocytotic activity in the ramified microglia. Possible physiological significance of this pinocytotic activity was indicated by the accumulation of various neurotransmitters/modulators: gamma-aminobutyric acid and vasoactive intestinal polypeptide (VIP). Ramified cells in these cultures have been previously noted to exhibit a constant and rapid pattern of motility, which was consistently observed here through time-lapse video recording; pinocytosis and rapid motility were shown to concur in individual cells. Based on their high intrinsic pinocytotic activity and pattern of cellular motility, the ramified microglia specifically are suggested to serve a constitutive function of fluid cleansing within the interstitial spaces of brain tissue.     

Variability in hand surface representations in areas 3b and 1 in adult owl and squirrel monkeys

Detailed microelectrode maps of the hand representation were derived in cortical areas 3b and 1 from a series of normal adult owl and squirrel monkeys. While overlap relationships were maintained, and all maps were internally topographic, many map features varied significantly when examined in detail. Variable features of the hand representations among different monkeys included a) the overall shapes and sizes of hand surface representations; b) the actual and proportional areas of representations of different skin surfaces and the cortical magnifications of representations of specific skin surfaces, which commonly varied severalfold in area 3b and manyfold in area 1; c) the topographic relationships among skin surface representations, with skin surfaces that were represented adjacently in some monkeys represented in locations many hundreds of microns apart in others; d) the internal orderliness of representations; e) the completeness of representations of the dorsal hand surfaces; and f) the skin surfaces represented along the borders of the hand representation. Owl monkey maps were, in general, internally more strictly topographic than squirrel monkey maps. In both species, area 3b was more strictly topographic and less variable than was area 1. The degree of individual variability revealed in these experiments is difficult to reconcile with the hypothesis that details of cortical maps are ontogenetically specified during a period in early life. Instead, we propose that differences in the details of cortical map structure are the consequence of individual differences in lifelong use of the hands. This conclusion is consistent with earlier studies of the consequences of peripheral nerve transection and digital amputation, which revealed that cortical maps are dynamically maintained and are alterable as a function of use or nerve injury in these monkeys (Merzenich et al., '83a,b, '84a; Merzenich, '86; Jenkins et al., '84; Jenkins and Merzenich, '87).     

Effect of CNTF on low-affinity NGF receptor expression by cultured neurons from different rat brain regions.

Our previous work indicated that in E14 embryonic rat spinal cord cultures ciliary neuronotrophic factor (CNTF) exerted (1) a survival-promoting effect on motor neurons and on a large population of unidentified neurons, and (2) a regulatory role on the expression of ChAT and low affinity NGF receptor (LNGFR) in a population of small/medium-sized neurons. In the present study, we examined the effect of CNTF on the expression of LNGFR in cultures of different regions from the E18 embryonic rat brain, namely cortex, septum, striatum, mesencephalon, hippocampus, brainstem, and cerebellum. The number of LNGFR-positive neurons (stained with the 192-IgG monoclonal antibody) was determined in untreated cultures and in cultures treated for 6 days (0-6) with human recombinant CNTF. To distinguish between effects on survival and on LNGFR expression, experiments were performed in which CNTF was administered only for the last 48 h of the culture (from days 4-6). LNGFR positive neurons were found in the cultures of all the regions examined. In each one of them, CNTF increased the number of LNGFR-positive neurons by three- to fourfold after 6 days of treatment. In the striatum, septum, mesencephalon, and cerebellum, the effect of CNTF was shown to be on the regulation of LNGFR expression and not on survival. In cultures from the cortex, hippocampus and brainstem, a survival-promoting role of CNTF could be demonstrated. The effect of CNTF was dose dependent, with half-maximal effects (ED50) achieved at 2-4.5 TU/ml for all the brain regions. Maximal effects were reached at 100-250 TU/ml. From these results, we conclude that (1) there exists a wide spectrum of CNTF-responsive neurons in the central nervous system, and (2) CNTF plays an important and widespread role in regulating the expression of the LNGFR in neurons.