EFFECT OF PARTIAL BRAIN ISCHAEMIA ON THE METABOLIC AND HAEMODYNAMIC RESPONSES TO HAEMORRHAGE HYPOTENSION MEASURED IN THE BRAIN AND SMALL INTESTINE Mira Mandelbaum-Livnat, Efrat Barbiro-Michaely and Avraham Mayevsky The Mina & Everard Goodman Faculty of Life-Sciences and The Gonda Multidisciplinary Brain Research Center Bar-Ilan University, Ramat-Gan, Israel ESCTAIC 23 rd Congress 2012 Timisoara, Romania October 4th 2012

Systemic level Tissue level cellular level Mitochondrial function preservation Anaerobic metabolism Cell injury Metabolic acidosis  Lactic acidosis Mitochondrial dysfunction Cell death  Circulatory blood volume  O 2 delivery Blood flow redistribution  Blood flow  Blood flow Sympathetic Nervous System activation Vasoconstriction  Cardiac output  Mean Arterial Pressure  Vessels resistance Autoregulation  Vessels resistance less vital organsvital organs Hemorrhage Ionic homeostasis disruption  ATP During hemorrhage blood is redistributed in favor of the vital organs and on the expense of the less vital organs. LDF-Laser Doppler Flowmetry

3 Simultaneous Real Time Monitoring of a Vital Organ - the Brain, and a Less Vital Organ - the Small Intestine, under Body Emergency Metabolic StatesBEMS) – a New Approach of Diagnostics Simultaneous Real Time Monitoring of a Vital Organ - the Brain, and a Less Vital Organ - the Small Intestine, under Body Emergency Metabolic States (BEMS) – a New Approach of Diagnostics Brain Vital Organ Small Intestine Less Vital Organ Blood Flow Redistribution under BEMS

4 Work hypothesis  During hemorrhage the body suffers from a decrease in oxygen delivery affecting primarily mitochondrial function.  During hemorrhage there is a redistribution of blood from the "less vital organs" (i.e. intestine and skin) to the life preserving circulation of the "vital organs" (i.e. brain and heart).  Monitoring of a “less vital organ” may early detect body emergency metabolic state occurring during hemorrhage.

5 Importance of research 50% of the patients with hemorrhagic shock die of substantial blood loss within an hour from the insult. Another 30% of deaths result from severe internal organ injury during the following minutes. Those who survive their initial injury are at a high risk of developing infection and multi organ failure, which can further lead to death. Early diagnosis of hemodynamic catastrophe and early resuscitation is most important in hemorrhagic shock in order to improve the final outcome.

6 Comparison of “vital organ” and a “less vital organ” may enable a better understanding of the process occurring on a daily basis in the clinic. The monitoring of less vital organ during hemorrhage has two important roles: 1. The early detection of the hemorrhage insult itself. 2. The early detection of resuscitation end point.

7 METHODS

The Multi-Site Multi-Parametric system for monitoring cerebral and intestinal blood flow and mitochondrial NADH

A1A1 A2A2 A BC B1B1 B2B2 C Brain Liver & kidney Testis D

10 The development of the monitoring model In accordance to recent studies reporting about homodynamic differences between two intestinal layers, namely the serosa and mucosa, following hemorrhage we carried out two protocols of short anoxia and epinephrine injection, in order to assure monitoring from both layers and to find out which intestinal location is better for intestinal monitoring.

11 Short anoxia Epinephrine I.V. injection Time (min) N2N2 N2N2 N 2 - death Start Time (min) N2N2 Epinephrine N 2 - death Start No significant differences between the serosa and mucosa were observed in any of the protocols. Therefore, we have decided to place the intestinal probe on the serosa, since it is less invasive and easier to manipulate.

12 Brain versus Intestine

13 Normotensive control 0123 Time (hour) N2N2 N 2 - death Start Start control N=4

14 Intestinal and Brain responses to Anoxia ) Start Stop Anoxia Time (sec) Small Intestinal Serosa Brain

15 Response of Intestine and Brain to Hypoxia StartStop Hypoxia ) ) Time (min) Small Intestinal Serosa Brain

16 Response of Intestine (gray) and Brain (black) to Hypercapnia

17 Responses of Intestine (gray) and Brain (black) to Hyperoxia

18 MAP ( mmHg ) TBF (%) NADH (%) REF (%) Intestinal and Brain responses to Epinephrine 10 µg/kg Small Intestinal Serosa Brain

19 MAP ( mmHg ) TBF (%) NADH (%) REF (%) ** Intestinal and Brain responses to Epinephrine 2-10 µg/kg Small Intestinal Serosa Brain

20 Anoxia (%) ***

21 Hypoxia (%) ***

22 Hypercapnia (%) ***

23 Correlations between NADH & TBF under Epinephrine 2-10  g/kg (I.V)

24 Hemorrhage Decreased circulatory blood volume Decreased tissue perfusion and O 2 delivery Oxygen demand exceeds oxygen supply Hemorrhagic shock

25 Bleeding down to 40 mmHg and maintenance 1234 Time (hour) N2N2 N 2 - death Start 0 Operation Resuscitation 15 min Sample protocol

26 Protocols Number of animals Protocol Development of the monitoring model 7Short anoxia 7Epinephrine I.V. injection The hemorrhage models 4Normotensive group 9Uncontrolled hypotension for 30 min 9Controlled hypotension for 15 min 12Controlled hypotension for 30 min 13Controlled hypotension for 60 min 4Partial cerebral ischemic control group 9Partial cerebral ischemia combined with hypotension

27 Uncontrolled hypotension Bleeding 0123 N2N2 Resuscitation N 2 - death Start 0123 Time (hour) 30 min Averaged amount of shed blood was calculated to be 12±3% of rat’s total blood volume. N=9

28 Controlled hypotension for 15 min Averaged amount of shed blood was calculated to be 31±2% of rat’s total blood volume. N=9

29 Bleeding and maintenance 0123 N2N2 Resuscitation N 2 - death Start 0123 Time (hour) 30 min Controlled hypotension for 30 min Averaged amount of shed blood was calculated to be 40±1.5% of rat’s total blood volume.

30 Graded Hemorrhage

31 Normal cerebral perfusion Partial cerebral ischemia Controlled hypotension under Normal cerebral perfusion versus Partial cerebral ischemia Comparison between various models of hypotension

32 Control Unilateral carotid Occlusion N=4

33 Bleeding after Unilateral carotid Occlusion N=9

Time (hour) N2N2 N 2 - death Start 0 Bilateral carotid occlusion 24 Operation Start control Partial cerebral ischemia - control group (no hemorrhage) Bilateral carotid occlusion (BCO) is an animal model of arteriosclerosis, which is considered to be the leading cause of mortality in industrialized countries.

Controlled hypotension for 15 min under partial cerebral ischemia Bleeding and maintenance Time (hour) N2N2 N 2 - death Start 0 Bilateral carotid occlusion 24 Operation Resuscitation 15 min

Brain Intestine The differences between the two models are manifested mainly by the cerebral responses. Comparison between the two organs in both experimental groups

TBFNADH Controlled 15 min Partial cerebral Ischemia The brain and intestinal responses to hemorrhage

Conclusions Under normal conditions  The early signs of the hemorrhage insult were detected in the intestine.  The initial deterioration of the intestine, following resuscitation, was not accompanied by a deterioration of MAP.  In all of the models the intestine suffered from irreversible damage, whereas the brain remained protected.  NADH responses to hemorrhage were higher in the intestine compared to the brain. Under partial cerebral ischemia  The response of the ischemic brain, to hemorrhage, was very similar to the intestinal response. The cerebral tissue was suffering from extensive reduction of blood flow.  The combination of partial cerebral ischemia and hemorrhagic hypotension blurs the differences between the brain and the small intestine.

39 Conclusions  The intestine may serve as a surrogate organ for monitoring under hemorrhagic insults, due to its capability for early detection of whole body deterioration.  The monitoring of mitochondrial NADH redox state can be used as an indicator of different hemorrhage stress severities. The application of the Multi-site Multi-parametric monitoring system is clearly advantageous under hemorrhagic hypotension.