In vivo temporal and spatial profile of leukocyte adhesion and migration after experimental traumatic brain injury in mice

Background: Leukocytes are believed to be involved in delayed cell death following traumatic brain injury (TBI). However, data demonstrating that blood-borne inflammatory cells are present in the injured brain prior to the onset of secondary brain damage have been inconclusive. We therefore investigated both the interaction between leukocytes and the cerebrovascular endothelium using in vivo imaging and the accumulation of leukocytes in the penumbra following experimentally induced TBI. Methods: Experimental TBI was induced in C57/Bl6 mice (n = 42) using the controlled cortical impact (CCI) injury model, and leukocyte-endothelium interactions (LEI) were quantified using both intravital fluorescence microscopy (IVM) of superficial vessels and 2-photon microscopy of cortical vessels for up to 14 h post-CCI. In a separate experimental group, leukocyte accumulation and secondary lesion expansion were analyzed in mice that were sacrificed 15 min, 2, 6, 12, 24, or 48 h after CCI (n = 48). Finally, leukocyte adhesion was blocked with anti-CD18 antibodies, and the effects on LEI and secondary lesion expansion were determined 16 (n = 12) and 24 h (n = 21), respectively, following TBI. Results: One hour after TBI leukocytes and leukocyte-platelet aggregates started to roll on the endothelium of pial venules, whereas no significant LEI were observed in pial arterioles or in sham-operated mice. With a delay of >4 h, leukocytes and aggregates did also firmly adhere to the venular endothelium. In deep cortical vessels (250 mu m) LEIs were much less pronounced. Transmigration of leukocytes into the brain parenchyma only became significant after the tissue became necrotic. Treatment with anti-CD18 antibodies reduced adhesion by 65\%; however, this treatment had no effect on secondary lesion expansion. Conclusions: LEI occurred primarily in pial venules, whereas little or no LEI occurred in arterioles or deep cortical vessels. Inhibiting LEI did not affect secondary lesion expansion. Importantly, the majority of migrating leukocytes entered the injured brain parenchyma only after the tissue became necrotic. Our results therefore suggest that neither intravascular leukocyte adhesion nor the migration of leukocytes into cerebral tissue play a significant role in the development of secondary lesion expansion following TBI

Vasopressin V1a Receptors Regulate Cerebral Aquaporin 1 after Traumatic Brain Injury

Brain edema formation contributes to secondary brain damage and unfavorable outcome after traumatic brain injury (TBI). Aquaporins (AQP), highly selective water channels, are involved in the formation of post-trauma brain edema; however, their regulation is largely unknown. Because vasopressin receptors are involved in AQP-mediated water transport in the kidney and inhibition of V$_{1a}$ receptors reduces post-trauma brain edema formation, we hypothesize that cerebral AQPs may be regulated by V$_{1a}$ receptors. Cerebral Aqp1 and Aqp4 messenger ribonucleic acid (mRNA) and AQP1 and AQP4 protein levels were quantified in wild-type and V$_{1a}$ receptor knockout (V$_{1a}$$^{-/-}$) mice before and 15 min, 1, 3, 6, 12, or 24 h after experimental TBI by controlled cortical impact. In non-traumatized mice, we found AQP1 and AQP4 expression in cortical neurons and astrocytes, respectively. Experimental TBI had no effect on Aqp4 mRNA or AQP4 protein expression, but increased Aqp1 mRNA (p < 0.05) and AQP1 protein expression (p < 0.05) in both hemispheres. The Aqp1 mRNA and AQP1 protein regulation was blunted in V$_{1a}$ receptor knockout mice. The V$_{1a}$ receptors regulate cerebral AQP1 expression after experimental TBI, thereby unraveling the molecular mechanism by which these receptors may mediate brain edema formation after TBI

Open Access

In vivo temporal and spatial profile of leukocyte adhesion and migration after experimental traumatic brain injury in mic

The role of bradykinin B1 and B2 receptors for secondary brain damage after traumatic brain injury in mice

Inflammatory mechanisms are known to contribute to the pathophysiology of traumatic brain injury (TBI). Since bradykinin is one of the first mediators activated during inflammation, we investigated the role of bradykinin and its receptors in posttraumatic secondary brain damage. We subjected wild-type (WT), B1-, and B2-receptor-knockout mice to controlled cortical impact (CCI) and analyzed tissue bradykinin as well as kinin receptor mRNA and protein expression up to 48 h thereafter. Brain edema, contusion volume, and functional outcome were assessed 24 h and 7 days after CCI. Tissue bradykinin was maximally increased 2 h after trauma (P<0.01 versus sham). Kinin B1 receptor mRNA was upregulated up to four-fold 24 h after CCI. Immunohistochemistry showed that B1 and B2 receptors were expressed in the brain and were significantly upregulated in the traumatic penumbra 1 to 24 h after CCI. B2R−/− mice had significantly less brain edema (−51% versus WT, 24 h; P<0.001), smaller contusion volumes (∼50% versus WT 24 h and 7 d after CCI; P<0.05), and better functional outcome 7 days after TBI as compared with WT mice (P<0.05). The present results show that bradykinin and its B2 receptors play a causal role for brain edema formation and cell death after TBI

Changes of cerebral blood flow during the secondary expansion of a cortical contusion assessed by <tex>^{14}C$</tex>-iodoantipyrine autoradiography in mice using a non-invasive protocol

Although changes of cerebral blood flow (CBF) in and around traumatic contusions are well documented, the role of CBF for the delayed death of neuronal cells in the traumatic penumbra ultimately resulting in secondary contusion expansion remains unclear. The aim of the current study was therefore to investigate the relationship between changes of CBF and progressive peri-contusional cell death following traumatic brain injury (TBI). CBF and contusion size were measured in C57Bl6 mice under continuous on-line monitoring of ETpCO2 before, and at 15 min and 24 h following controlled cortical impact by C-14-iodoantipyrine autoradiography (IAP-AR; n = 5-6 per group) and by Nissl staining, respectively. Contused and ischemic (CBF < 10%) tissue volumes were calculated and compared over time. Cortical CBF in not injured mice varied between 69 and 93 mL/100mg/min depending on the anatomical location. Fifteen minutes after trauma, CBF decreased in the whole brain by similar to 50% (39 +/- 18 mL/100mg/min; p < 0.05), except in contused tissue where it fell by more than 90% (3 +/- 2 mL/100mg/min; p < 0.001). Within 24 h after TBI, CBF recovered to normal values in all brain areas except the contusion where it remained reduced by more than 90% (p < 0.001). Contusion volume expanded from 24.9 to 35.5 mm3 (p < 0.01) from 15 min to 24 h after trauma (+43%), whereas the area of severe ischemia (CBF < 10%) showed only a minimal (+13%) and not significant increase (22.3 to 25.1 mm(3)). The current data therefore suggest that the delayed secondary expansion of a cortical contusion following traumatic brain injury may not be caused by a reduction of CBF alone