Abstract
Global ischaemia during cardiac surgery, brain surgery and stroke frequently leads to neuronal injury and lasting neuropsychological dysfunction. Experimental preconditioning provides beneficial cerebral outcomes in these conditions. In this study, the neuroprotective effects of low-dose GYKI-52466 prophylactic preconditioning were evaluated by histological, behavioural, and electrophysiological paradigms in a rat model of hypoxia-ischaemia (HI). Male Sprague Dawley rats (postnatal day 26) were subcutaneously administered with saline or GYKI-52466 (GYKI; 3 mg/kg, 90 min; 1 mg/kg, twice in 120 min; or 0.5 mg/kg, thrice in 180 min) prior to left common carotid artery occlusion. Animals were allowed to recover for 2 hrs, and then placed in a hypoxia chamber (8% O2/92% N2; 33 ± 1°C) for 1 hr. A sham surgery group received saline without HI. Histological assessment was carried out on days 14, and 90 and sensorimotor behavioural tests were performed before surgery and at 1, 7, 14 and 90 days post-HI. Low-dose GYKI-52466 preconditioning significantly reduced infarct volume and ventricular enlargement relative to saline treated controls at day 14 after HI. On day 90, tissue loss was significantly reduced by GYKI 3 mg/kg compared to saline. Foot-faults, paw use asymmetry, and postural reflex scores were significantly improved in all GYKI treatment groups.
Hippocampal synaptic plasticity is considered a cellular basis of learning and memory. Therefore, electrophysiological assessment was performed on days 14 and 90 to assess hippocampal CA1 long-term potentiation (LTP) and long-term depression (LTD). The contralateral and, where possible, ipsilateral hippocampal slices were prepared from animals preconditioned with saline or GYKI-52466 (3 mg/kg), and population spikes and field EPSPs recorded in balanced (2 mM Ca2+ and 2 mM Mg2+) artificial cerebrospinal fluid at 31 ± 1°C. GYKI-52466 preconditioning significantly reversed the HI-induced suppression of neuronal excitability and synaptic strength in both contralateral and ipsilateral hippocampi on both test days. After HI, LTP was significantly reduced in the contralateral and entirely absent in ipsilateral hippocampi, but LTD was unchanged. In ischaemic animals preconditioned with GYKI-52466, LTP was readily induced on both sides. Previously, it has been hypothesised that GYKI-52466 can trigger strong, presumably metabotropic, protection against seizures at exceptionally low doses. To this end, we determined the brain concentrations of GYKI-52466 following subcutaneous administration of 3 and 20 mg/kg of GYKI-52466 in control and brain damaged rats. A low brain concentration of 0.56 μM GYKI-52466 was observed with 3 mg/kg compared to 10.7 μM with 20 mg/kg at 90 minutes post drug administration. GYKI-52466 at 20 mg/kg produced severe ataxia up to 90 min. Furthermore, in ischaemic animals, there was no evidence of a 'surge' in brain GYKI-52466 concentrations at the infarct, confirming the integrity of blood-brain barrier.
These results provide evidence of long-term functional neuroprotection by GYKI-52466 at doses well below, and at pre-administration intervals well-beyond previous studies, and suggest that a classical blockade of ionotropic AMPA receptors does not underlie its neuroprotective effects. Low-dose GYKI-52466 preconditioning represents a novel, prophylactic strategy for neuroprotection in a field almost devoid of effective pharmaceuticals.