In CA1 pyramidal neurones a slow afterhyperpolarisation current (IsAHP) regulates action potential firing frequency. Exogenous kainate application (10-100 nM) inhibits IsAHP leading to an increase in excitability. This effect is mediated by a metabotropic action of postsynaptic kainate receptors (KARs), presumably containing GluR6 subunits, on CA1 pyramidal cells (Melyan et al. 2002). We investigated whether synaptic activation of these receptors can evoke inhibition of IsAHP similar to that produced by kainate application.
Hippocampal slices were obtained from humanely killed 14- to 21-day-old rats. Whole-cell recordings using KMeSO4-based solution were made from CA1 pyramidal cells. Voltage steps (60 mV for 80 ms) were applied every 20 s to generate IsAHP. The effect of synaptically released glutamate was assessed by comparing IsAHPs induced before and after a train of stimuli delivered via a stimulating electrode (Heuss et al. 1999). Data are expressed as means ± S.E.M.
Trains of synaptic stimuli (5 pulses, 100 Hz) previously shown to activate KARs (Min et al. 1999) produced 33 ± 1 % inhibition of IsAHP amplitude (n = 8). In order to isolate KAR responses, experiments were carried out in the presence of a cocktail of antagonists to block NMDAR (100 µM DL-AP5), AMPAR (100 µM GYKI52466), mGluR (1 mM MCPG and 250 µM MSOP), GABAAR (100 µM picrotoxin), GABABR (200 µM 2-OH-saclofen), muscarinic AChR (1 µM atropine sulphate), opioid (10 µM naloxone), cannabinoid CB1 (2 µM AM 251) and adenosine receptors (0.1 µM DPCPX). The IsAHP inhibition was not reversible within 30 min, similar to observations with kainate application. The IsAHP inhibition was effectively blocked by prior application of 20 µM CNQX (n = 6) confirming that it was mediated by kainate receptor activation. The glutamate uptake blocker TBOA (50-100 µM) increased the effect of synaptic stimulation on IsAHP to 50 ± 2 % inhibition (n = 8). Increasing the number of stimuli to 10 or 20 also increased IsAHP inhibition to 38 ± 7 % (n = 4) and 54 ± 2 % (n = 4), respectively, whereas a single stimulus had no effect (n = 3).
We conclude that synaptically released glutamate acting on kainate receptors can reduce IsAHP in CA1 pyramidal neurons indicating a direct role for kainate receptors in regulation of CA1 pyramidal cell excitability.
The work is supported by The Wellcome Trust.