Localized Hypothermia Aggravates Bleeding in the Collagenase Model of Intracerebral Hemorrhage

Abstract

Animal studies testing whether therapeutic hypothermia is neuroprotective after intracerebral hemorrhage (ICH) have been inconclusive. In rodents, ICH is often produced in the striatum by infusing collagenase, which causes prolonged hemorrhaging from multiple vessels. Our previous data shows that this bleeding (hematoma) is worsened by systemic hypothermia given soon after collagenase infusion. In this study we hypothesized that localized brain hypothermia would also aggravate bleeding in this model (0.2 U of collagenase in 1.2 μL of saline). We also evaluated cooling after intrastriatal thrombin infusion (1 U in 30 μL of saline)-a simplified model of ICH thought to cause bleeding. Focal hypothermia was achieved by flushing cold water through an implanted cooling device attached to the skull underneath the temporalis muscle of adult rats. Previous work and data at this time shows this method cools the striatum to ∼33°C, whereas the body remains normothermic. In comparison to normothermic groups, cooling significantly worsened bleeding when instituted at 6 hours (∼94 vs. 42 μL, p=0.018) and 12 hours (79 vs. 61 μL, p=0.042) post-ICH (24-hour survival), but not after a 24-hour delay (36-hour survival). Rats were cooled until euthanasia when hematoma size was determined by a hemoglobin-based spectrophotometry assay. Cooling did not influence cerebral blood volume after just saline or thrombin infusion. The latter is explained by the fact that thrombin did not cause bleeding beyond that caused by saline infusion. In summary, local hypothermia significantly aggravates bleeding many hours after collagenase infusion suggesting that bleeding may have confounded earlier studies with hypothermia. Furthermore, these findings serve as a cautionary note on using cooling even many hours after cerebral bleeding.

Acetazolamide Mitigates Intracranial Pressure Spikes Without Affecting Functional Outcome After Experimental Hemorrhagic Stroke

“…We have repeatedly used this technique to monitor minute-by-minute ICP changes in freely moving rats over several days [7, 13–16]. In addition, we tested whether acetazolamide affects hematoma formation because it can acutely increase cerebral blood flow [17], and early interventions can aggravate bleeding after collagenase-induced ICH [18]. Finally, we assessed effects of acetazolamide on behavioral and histological outcome.…”

Examining potential side effects of therapeutic hypothermia in experimental intracerebral hemorrhage

“…Although not part of our initial hypothesis, the most interesting and concerning result of this study is the influence of TH on bleeding post-treatment, since the hematoma volume is a critical predictor of outcome and survival. 49,50 Previous studies show that TH applied too early can aggravate bleeding, 22,25 which we successfully avoided by delaying treatment, as we have previously done. 24 As there were no group differences in hematoma volume at 1 and 3 days post-ICH, the increase seen on day 7 is likely due to re-bleeding either during rewarming or over the following days.…”

“…for hemoglobin levels after ICH the average standard deviation (S.D.) was $15 after ICH, 25 and for non-heme iron the SD was 1.2 on day 3 after ICH 38 ). From that we roughly obtain estimates for effect sizes and variability with the goal of having 80% power to detect those effects.…”

“…32 The greater bleeding is essential as it can be an important complication of using TH to treat ICH. 22,25 Materials and methods…”

Infusion of Cold Saline into the Carotid Artery Does Not Affect Outcome After Intrastriatal Hemorrhage

“…TH can affect hemostasis by inhibiting clotting factors and altering platelet function (Alam et al , 2012 ). We showed that prolonged hypothermia (HYPO) aggravates bleeding in the collagenase ICH model (John et al , 2015 ). Therefore, depth, duration, and onset of TH must be considered when TH is used in populations with a high risk of hemorrhage.…”