Atsuo Miyakawa

In vivo calcium imaging of cerebral cortex in hypoxia-ischemia followed by developmental stage-specific injury in rats

The parietal area is a part of the cortex that is vulnerable in the rat to hypoxia-ischemia (HI) within the early postnatal period. To investigate the localizing mechanism of this cortical injury, we spatiotemporally detected the cortical intracellular calcium changes, as revealed by a calcium-sensitive fluorescence dye, Rhod 2-AM, during 1h of HI on postnatal days 7-21 in vivo. The calcium level rose to different levels at different cortical points in all animals within the first 20 min. Over the whole cortical area in the camera field, the changes in three groups significantly differed across time at 30 and 60 min, and a chronic increase appeared at days 7-8. After 3h of reperfusion, microtubule-associated protein 2 (MAP-2) immunoreactivity confirmed that parietal injury was more serious at day 7, whereas the imaging of calcium distribution did not segregate the injured and uninjured areas. Our in vivo findings in the whole brain structure indicate that the age-specific vulnerability of the parietal cortex injury is affected indirectly by the chronic increase in the late HI phase in the early postnatal period, suggesting that each cortical area differs postnatally with respect to the development of calcium regulation and signal transduction involving neural cell death and/or survival.

Imaging of two types of calcium-transients induced by neuronal excitations in the rat hippocampal slice

Optical imaging of the intrinsic signal has been used to visualize functional structures. This signal induced by neural activation is related to concentration changes of deoxy-hemoglobin (deoxy-Hb), oxy-hemoglobin (oxy-Hb), blood volume (BV), and change in light scattering (LS). In this study we investigated spatial patterns of orientation columns derived from these different components in anesthetized and awake cat cortices. We recorded the intrinsic signal at 540, 570, and 620 nm simultaneously by using three CCD cameras and decomposed it into the above-mentioned components, A flashed grating or moving grating was used for visual stimulation. The average response evoked by two orthogonal gratings indicated the global change (GC) for each component and a difference image obtained from the two gratings indicated the local difference (LD) of each component. The LD of BV showed an identical columnar pattern with that of deoxy-Hb. These small laocal changes were superimposed on a larger global changes across the cortex, with increasing GC of BV, and deoxy-Hb. These results suggest that, as well as deoxy-Hb, neural activation induces a large increase in BV both in active and less active columns, but the amount is larger in active columns. We hypothesize that there is a fine control mechanism of blood flow at the columnar level. Analysis of oxy-Hb, LS and differences in awake and anesthetized condition will be also discussed.

1232 Calcium macroimaging of cortex of young rats in hypoxia-ischemic condition

TAKESHI KIHARA’, SHUN SHIMOHAMA’, HIDEYUKI SAWADAl, JUN KIMURA’, TOSHIAKI KUME2, AKINORI AKAIKE2 The effects of nicotinic acetylcholine receptor-related substances on beta amyloid (AD) toxicity were examined using primary cultures of rat cortical neurons. The number of viable neurons decreased when cultures were exposed to A/3. Incubating the cultures with nicotine markedly reduced AD toxicity. This nicotine-induced protection was blocked by a selective a7 nicotinic receptor antagonist, alpha-bungarotoxin ((r-BTX). Furthermore, dimethoxybenzylidene anabaseine (DMXB), a selective a7 receptor agonist, decreased the dead cells induced by Ap. On the other hand, dihydro-beta-erythroidine (DDE), a neuronal CNS receptor antagonist, blocked the protective effect of nicotine against AP toxicity. And cytisine, thought to be a neuronal CNS receptor agonist, inhibited AD toxicity. These results suggest that nicotinic receptor stimulation protect neurons from degeneration induced by A/3 and that both ~7 and neuronal CNS receptors play an important role in the mechanism of the neuroprotection.