Interleukin-1β Blocks the Effect of BDNF on Rat Brain Mitochondria

Anthony Markham1, Rasneer Bains1, Michael Spedding2, Paul Franklin1
1University of Sunderland, SUNDERLAND SR1 3SD, United Kingdom, 2Institut de Recherches Internationales Servier, 31, Rue du Pont, Neuilly sur Seine, France

Interleukin-1β (IL-1β) is a pro-inflammatory cytokine involved in modulation of inflammation and stress responses in the brain. Administration of IL-1β impairs hippocampal memory formation (Song et al., 2004) and also long-term potentiation (LTP: Bellinger et al., 1993), the long-term increase in synaptic transmission thought to underlie memory formation. Brain derived neurotrophic factor (BDNF) plays a vital role in the process of LTP (Jankowski & Patterson, 1999) and has been shown to modulate brain mitochondrial function, the key organelle in coupling neuronal plasticity to synaptic energy expenditure (Markham et al., 2004). We therefore wished to investigate the effect of IL-1β on BDNF induced mitochondrial protection.

Forebrains from female Wistar rats (250-300g) were rapidly removed and placed in isolation buffer containing 220mM mannitol, 60mM sucrose, 5mM Tris-HCl, 0.5mM EGTA and 1mg ml-1 bovine serum albumin (BSA; fatty acid free) pH 7.4. Homogenates were centrifuged at 2,000 rpm for 6 min at 0-4°C and the resulting supernatant decanted off and spun for 8 min at 10,000 rpm. The pellet produced was resuspended in 9 ml of buffer and aliquots layered onto 10 ml ice-cold Percoll solution containing 250mM sucrose, 5mM Tris-HCl, 0.1mM EGTA and 18% (w/v) Percoll, pH 7.4. The resulting density gradient was centrifuged at 10,000 rpm for 45 min. A loose mitochondrial pellet, free from most contamination, was formed at the bottom of the tube with a synaptosomal fraction forming a separate layer at the top of the tube. The two layers were then isolated and centrifuged at 10,000 rpm in isolation buffer minus EGTA (incubation buffer) to remove the Percoll.

Oxygen consumption was measured polarographically (Sweetman & Weetman, 1972) using a Clark-type oxygen electrode (Rank Bros, Bottisham, UK). Respiratory studies were performed after the method of Markham et al., (2004) and recorded on a Kipp and Zonen Flat bed recorder. Calculating the RCI (Respiratory Control Index; a measure of the efficiency of respiratory coupling) assessed mitochondrial integrity. Data were analysed using a one-way ANOVA followed by Dunnett’s post-test.

BDNF (333 ng ml-1) significantly increased brain mitochondrial RCI (6.89 ± 0.19) compared to control (5.22 ± 0.23; P<0.01; n = 3). IL-1β alone (333 ng ml-1) had no significant effect on RCI (5.33 ± 0.17 vs. 5.22 ± 0.23; P>0.5), but significantly decreased the effect of BDNF (6.89 ± 0.19 vs. 5.20 ± 0.33; P<0.01; n = 3).

In addition to blocking the effect of BDNF on LTP, IL-1β blocks the increase in mitochondrial respiratory efficiency brought about by BDNF. The results indicate the possible actions of different pathways converging on the mitochondrion to change mitochondrial respiratory efficiency.

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