Fig. 1. Caspase-3 activation after volatile agent exposure. (  A ) Low-power (10× objective) view of the dorsal hippocampus and overlying cortex after 6 h of isoflurane exposure. The channel acquiring signal from the Alexa-fluor 488 is shown. The  green cells are activated caspase-3–positive cells in CA-1 and the overlying cortex. The  left edge of the slide is medial, and the  top edge isdorsal. (  B ) Low-power (10× objective) view of the cingulate cortex 6 h of isoflurane exposure. The merged channel shows both the activated caspase-3–positive cells (  green ) and NeuN-positive cells (  red ). Cells with an  orange to yellow hue are positive for both signals and represent mature neurons that are caspase-positive. (  C ) High-power (63× oil objective) view if the cortex overlying CA-1 after 6 h of isoflurane exposure, merged channel. Several activated caspase-3–positive cells are seen, and these have a morphology suggestive of mature neurons that have degenerated. There is also evidence of nodular staining along dendritic branches throughout the image. (  D ) Low-power view (with high-power inset) of subiculum at level of dorsal hippocampus after 3 h of desflurane exposure. Activated caspase-3–positive cells are seen in the subiculum on the low-power view. Very few caspase-positive cells are seen in the overlying retrosplenial cortex (compare to  Panel A ). The high-power view shows that some of the caspase-positive cells are colocalized with NeuN-positive cells. In addition, a blood vessel with stained nucleated red blood cells is visible on the  right-hand side of the inset. (  E ) Low-power (10× objective with 2× zoom) view of the retrosplenial cortex and medial portion of CA1 after 3 h of isoflurane exposure. A few caspase-3–positive cells are seen after 3 h of isoflurane in the retrosplenial cortex but far fewer than after 6 h of exposure. Activated caspase-3–positive cells are also present in corpus callosum and the fasciola ceruneum. (  F ) High-power (63× oil objective) view of the superficial layers of the primary motor cortex at the level of the dorsal striatum after 3 h of sevoflurane exposure. A dendritic caspase-3–positive cell is seen in detail as well as some other slightly out of the plane of focus. The cell shown did not colocalize with the NeuN signal in any of the 10 optical planes in the z-stack, indicating this cell is probably an astrocyte or reactive microglial cell in which caspase-3 has been activated. 

Fig. 1. Caspase-3 activation after volatile agent exposure. (  A ) Low-power (10× objective) view of the dorsal hippocampus and overlying cortex after 6 h of isoflurane exposure. The channel acquiring signal from the Alexa-fluor 488 is shown. The  green cells are activated caspase-3–positive cells in CA-1 and the overlying cortex. The  left edge of the slide is medial, and the  top edge isdorsal. (  B ) Low-power (10× objective) view of the cingulate cortex 6 h of isoflurane exposure. The merged channel shows both the activated caspase-3–positive cells (  green ) and NeuN-positive cells (  red ). Cells with an  orange to yellow hue are positive for both signals and represent mature neurons that are caspase-positive. (  C ) High-power (63× oil objective) view if the cortex overlying CA-1 after 6 h of isoflurane exposure, merged channel. Several activated caspase-3–positive cells are seen, and these have a morphology suggestive of mature neurons that have degenerated. There is also evidence of nodular staining along dendritic branches throughout the image. (  D ) Low-power view (with high-power inset) of subiculum at level of dorsal hippocampus after 3 h of desflurane exposure. Activated caspase-3–positive cells are seen in the subiculum on the low-power view. Very few caspase-positive cells are seen in the overlying retrosplenial cortex (compare to  Panel A ). The high-power view shows that some of the caspase-positive cells are colocalized with NeuN-positive cells. In addition, a blood vessel with stained nucleated red blood cells is visible on the  right-hand side of the inset. (  E ) Low-power (10× objective with 2× zoom) view of the retrosplenial cortex and medial portion of CA1 after 3 h of isoflurane exposure. A few caspase-3–positive cells are seen after 3 h of isoflurane in the retrosplenial cortex but far fewer than after 6 h of exposure. Activated caspase-3–positive cells are also present in corpus callosum and the fasciola ceruneum. (  F ) High-power (63× oil objective) view of the superficial layers of the primary motor cortex at the level of the dorsal striatum after 3 h of sevoflurane exposure. A dendritic caspase-3–positive cell is seen in detail as well as some other slightly out of the plane of focus. The cell shown did not colocalize with the NeuN signal in any of the 10 optical planes in the z-stack, indicating this cell is probably an astrocyte or reactive microglial cell in which caspase-3 has been activated. 

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