Source Paper
Interleukin-1β Immunoreactivity and Microglia Are Enhanced in the Rat Hippocampus by Focal Kainate Application: Functional Evidence for Enhancement of Electrographic Seizures
Annamaria Vezzani, Mirko Conti, Ada De Luigi, Teresa Ravizza, Daniela Moneta et al.
Journal of Neuroscience • 1999
IL-1β Functional Effect on Seizure Duration
Objective: Measure the effect of exogenous human recombinant IL-1β on kainate-induced seizure duration and determine if IL-1β receptor antagonist and NMDA receptor antagonist can block this effect
Gather these items before starting the experiment. Check off items as you prepare.
Equipment1
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Materials5
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Software1
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Protocol Steps
Baseline assessment and animal preparation
Prepare rats for intrahippocampal injection procedure
Note: Unilateral intrahippocampal injection approach used
View evidence from paper
“unilateral intrahippocampal application of 0.19 nmol of kainate”
Intrahippocampal injection of human recombinant IL-1β
Inject 1 nanogram of hrIL-1β intrahippocampally 10 minutes before kainate injection
Note: Timing is critical - must be 10 minutes prior to kainate
View evidence from paper
“One nanogram of (hr)IL-1β intrahippocampally injected 10 min before kainate”
Intrahippocampal injection of kainate
Inject 0.19 nmol of kainic acid intrahippocampally to induce seizures
Note: This is the primary convulsant stimulus
View evidence from paper
“Three and 24 hr after unilateral intrahippocampal application of 0.19 nmol of kainate”
EEG recording and seizure monitoring
Record electroencephalographic activity to measure seizure duration and electrographic seizures
Note: Seizure duration is the primary outcome measure
View evidence from paper
“we measured the effect of human recombinant (hr)IL-1β on seizure activity as one marker of the response to kainate”
Blocking experiment with IL-1β receptor antagonist
Coinjection of 1 μg hrIL-1β receptor antagonist with hrIL-1β to determine if IL-1β effects are blocked
Note: Coinjection approach used to test blocking effects
View evidence from paper
“This effect was blocked by coinjection of 1 μg (hr)IL-1β receptor antagonist”
Blocking experiment with NMDA receptor antagonist
Coinjection of 0.1 ng of 3-((+)-2-carboxypiperazin-4-yl)-propyl-1-phosphonate with hrIL-1β to determine if NMDA receptor antagonism blocks IL-1β effects
Note: Selective NMDA receptor antagonist used
View evidence from paper
“or 0.1 ng of 3-((+)-2-carboxypiperazin-4-yl)-propyl-1-phosphonate, selective antagonists of IL-1β and NMDA receptors, respectively”
Tissue collection for immunocytochemistry and ELISA
Collect hippocampal tissue at 3 and 24 hours after kainate injection for analysis of IL-1β immunoreactivity and concentration
Note: Both injected and contralateral hippocampi collected
View evidence from paper
“Three and 24 hr after unilateral intrahippocampal application of 0.19 nmol of kainate, IL-1β immunoreactivity was enhanced”