The neurotoxic effect of isoflurane on age-defined neurons generated from tertiary dentate matrix in mice

Abstract

Introduction: Recent animal studies showed that isoflurane exposure may lead to the disturbance of hippocampal neurogenesis and later cognitive impairment. However, much less is known about the effect of isoflurane exposure on the neurons generated form tertiary dentate matrix, even though a great increase of granule cell population during the infantile period is principally derived from this area.

Methods: To label the new cells originated from the tertiary dentate matrix, the mice were injected with BrdU on postnatal day 6 (P6). Then, the mice were exposed to isoflurane for 4 hr at 1, 8, 21, and 42 days after BrdU injection, and the brains were collected 24 hr later. The loss of newly generated cells/neurons with different developmental stage was assessed by BrdU, BrdU + DCX, BrdU + NeuN, or BrdU + Prox-1 staining, respectively.

Results: We found that the isoflurane exposure significantly decreased the numbers of nascent cells (1 day old) and mature neurons (42 days old), but had no effect on the immature (8 days old) and early mature neurons (8 and 21 days old, respectively).

Conclusion: The results suggested isoflurane exposure exerts the neurotoxic effects on the tertiary dentate matrix-originated cells with an age-defined pattern in mice, which partly explain the cognitive impairment resulting from isoflurane exposure to the young brain.

Keywords: anesthetics; isoflurane; neurotoxicity; tertiary dentate matrix.

Figure 1

BrdU staining: the effect of isoflurane on the nascent cells. (a) There are significantly more BrdU cells in the ML, GC of DG in the control group than isoflurane‐treated group. (b) Quantitative analysis of the number of BrdU‐positive cells between the two groups. All data are presented as mean ± SEM.n = 5 for each group. #p ˂ 0.01 versus control. Scale bar = 100 µm

Figure 2

Double‐labeling staining of BrdU + DCX: the effect of isoflurane on the immature neurons. (a) Representative double immunofluorescence images of BrdU + DCX‐positive cells in the DG. (b) Quantitative analyses of BrdU + DCX‐positive cells in the DG show that there is no significant difference on the number of double‐labeling cells between the two groups. All data are presented as mean ± SEM.n = 5 for each group.*p ˂ 0.05 versus control. Scale bar = 100 µm

Figure 3

Double‐labeling staining of BrdU + Prox‐1 and BrdU + NeuN: the effect of isoflurane on the early mature neurons. a and c: Representative double immunofluorescence staining images of BrdU + Prox‐1‐ and BrdU + NeuN‐positive cells in DG. b and d: Quantitative analyses of double‐labeling positive cells do not find any difference on the number of double‐labeling cells between the two groups. All data are presented as mean ± SEM.n = 5 for each group. *p ˂ 0.05 versus control. Scale bar = 100 µm

Figure 4

Double‐labeling staining of BrdU + Prox‐1 and BrdU + NeuN: the effect of isoflurane on the mature neurons a and c: Representative double immunofluorescence staining images of BrdU + Prox‐1‐ and BrdU + NeuN‐positive cells in the DG. b and d: Quantitative analyses of double‐labeling positive cells show that the number of double‐labeling cells reduced significantly in the isoflurane‐treated group compared with the control group. All data are presented as mean ± SEM.n = 5 for each group.*p ˂ 0.05 and #p < .01 versus control. Scale bar = 100 µm