This series of experiments investigated whether the NMDA receptor antagonist D-2-amino-5-phosphonopentanoate (D-AP5) could induce impairments of spatial learning across a dose range comparable to its impairment of hippocampal long-term potentiation (LTP) in vivo. Estimations of the extracellular concentration of D-AP5 in hippocampus using microdialysis were also made to compare whether these impairments occur at concentrations similar to those required to impair LTP in the in vitro hippocampal slice. Rats were chronically infused with D-AP5 into the lateral ventricle at a range of concentrations (0–50 mM) via osmotic minipumps. They were first trained to find and escape onto a hidden platform in an open-field water maze task. After the behavioral learning, they were anesthetized with urethane and an attempt was made to evoke and monitor hippocampal LTP. Extracellular samples of D-AP5 in hippocampus were then taken using microdialysis, and finally, the animals were killed and tissue samples dissected. The microdialysis and tissue samples were analyzed for D-AP5 content using HPLC with fluorescence detection. The results established, first, that D-AP5 impairs spatial learning in a linear dose-dependent manner, highly correlated with its corresponding impairment of hippocampal LTP in vivo. No concentration of D-AP5 was observed to block LTP without affecting learning. Second, the microdialysis estimates indicated that, subject to certain assumptions, D-AP5 causes these impairments at extracellular concentrations comparable to those that impair LTP in vitro. Third, comparison of the whole tissue and microdialysis samples revealed a concentration ratio of approximately 30:1, indicating that 97% of the intracerebral D-AP5 is inaccessible to the dialysis probes. Infusion of 20 mM EGTA was found to cause a sevenfold increase in D-AP5 in the dialysis perfusates, suggesting that at least part of the inaccessible D-AP5 is trapped by a calcium-dependent mechanism. Two further behavioral control studies indicated that the D-AP5-induced impairment of spatial learning is unlikely to be secondary to a drug- induced motor disturbance, and that the performance of the D-AP5 group whose concentration was just sufficient to block hippocampal LTP completely was statistically indistinguishable from that of a group of rats with bilateral hippocampal lesions induced by ibotenic acid. Taken together, these findings offer support for the hypothesis that activation of NMDA receptors is necessary for certain kinds of learning.
Objective: To investigate whether the NMDA receptor antagonist D-AP5 impairs spatial learning in a dose-dependent manner comparable to its impairment of hippocampal LTP in vivo, and to determine the extracellular concentration of D-AP5 in hippocampus using microdialysis
This is a Morris Water Maze protocol using rat as the model organism. The procedure involves 9 procedural steps, 4 equipment items, 4 materials. Extracted from a 1992 paper published in Journal of Neuroscience.
Model and subjects
rat • Not specified • unknown • Not specified • Not specified
Study window
Estimated timing pending
Core workflow
Chronic D-AP5 infusion setup • Morris Water Maze training • Anesthesia and LTP recording preparation
Primary readouts
Key equipment and reagents
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Rats were chronically infused with D-AP5 into the lateral ventricle using osmotic minipumps at a range of concentrations from 0 to 50 mM
Note: Multiple concentration groups were tested to establish dose-response relationship
“Rats were chronically infused with D-AP5 into the lateral ventricle at a range of concentrations (0–50 mM) via osmotic minipumps”
Rats were trained to find and escape onto a hidden platform in an open-field water maze task to assess spatial learning
Note: This behavioral learning task was performed while D-AP5 was being infused
“They were first trained to find and escape onto a hidden platform in an open-field water maze task”
After behavioral learning, rats were anesthetized with urethane in preparation for hippocampal LTP recording
Note: Anesthesia was necessary for invasive LTP recording procedures
“After the behavioral learning, they were anesthetized with urethane and an attempt was made to evoke and monitor hippocampal LTP”
Hippocampal LTP was evoked and monitored in anesthetized rats to assess the electrophysiological effects of D-AP5
Note: LTP measurements were compared across different D-AP5 concentration groups
“an attempt was made to evoke and monitor hippocampal LTP”
Extracellular samples of D-AP5 in hippocampus were collected using microdialysis to determine the concentration of drug accessible to neural tissue
Note: Microdialysis was performed after LTP recording while animals were still anesthetized
“Extracellular samples of D-AP5 in hippocampus were then taken using microdialysis”
Animals were killed and tissue samples were dissected from hippocampus for analysis of D-AP5 content
Note: Tissue samples were collected after microdialysis sampling
“finally, the animals were killed and tissue samples dissected”
Microdialysis and tissue samples were analyzed for D-AP5 content using HPLC with fluorescence detection
Note: This analysis allowed comparison of extracellular versus total tissue D-AP5 concentrations
“The microdialysis and tissue samples were analyzed for D-AP5 content using HPLC with fluorescence detection”
In a separate experiment, 20 mM EGTA was infused to test whether calcium-dependent mechanisms sequester D-AP5
Note: EGTA infusion resulted in a sevenfold increase in D-AP5 in dialysis perfusates, suggesting calcium-dependent sequestration
“Infusion of 20 mM EGTA was found to cause a sevenfold increase in D-AP5 in the dialysis perfusates”
Two additional behavioral control studies were conducted to rule out motor disturbances and to compare D-AP5 effects with hippocampal lesion effects
Note: One control group received bilateral hippocampal lesions induced by ibotenic acid for comparison
“Two further behavioral control studies indicated that the D-AP5-induced impairment of spatial learning is unlikely to be secondary to a drug-induced motor disturbance”
This section explains what the experiment is doing, which readouts matter, what the data artifacts usually look like, and how the analysis should flow from raw capture to reported result.
To investigate whether the NMDA receptor antagonist D-AP5 impairs spatial learning in a dose-dependent manner comparable to its impairment of hippocampal LTP in vivo, and to determine the extracellular concentration of D-AP5 in hippocampus using microdialysis
Objective
To investigate whether the NMDA receptor antagonist D-AP5 impairs spatial learning in a dose-dependent manner comparable to its impairment of hippocampal LTP in vivo, and to determine the extracellular concentration of D-AP5 in hippocampus using microdialysis
Subjects
From paperrat • Not specified • unknown • Not specified • Not specified
Cohort notes
From paperSome rats received bilateral hippocampal lesions induced by ibotenic acid as a control group
Chronic D-AP5 infusion setup (Not specified)
Morris Water Maze training (Not specified)
Anesthesia and LTP recording preparation (Not specified)
Hippocampal LTP evocation and monitoring (Not specified)
Spatial learning performance in Morris Water Maze (escape latency, path length, or similar metrics)
From paperCorrelation analysis was performed between D-AP5-induced impairment of spatial learning and corresponding impairment of hippocampal LTP in vivo.
Artifact type
Representative image panels with region or marker comparisons
Comparison focus
Compare staining intensity, structure, or cell counts across matched conditions
Hippocampal LTP magnitude and induction
From paperCorrelation analysis was performed between D-AP5-induced impairment of spatial learning and corresponding impairment of hippocampal LTP in vivo.
Artifact type
Representative image panels with region or marker comparisons
Comparison focus
Compare staining intensity, structure, or cell counts across matched conditions
Extracellular D-AP5 concentration in hippocampus via microdialysis
From paperCorrelation analysis was performed between D-AP5-induced impairment of spatial learning and corresponding impairment of hippocampal LTP in vivo.
Artifact type
Representative image panels with region or marker comparisons
Comparison focus
Compare staining intensity, structure, or cell counts across matched conditions
Total tissue D-AP5 concentration
From paperCorrelation analysis was performed between D-AP5-induced impairment of spatial learning and corresponding impairment of hippocampal LTP in vivo.
Artifact type
Representative image panels with region or marker comparisons
Comparison focus
Compare staining intensity, structure, or cell counts across matched conditions
Spatial learning performance in Morris Water Maze (escape latency, path length, or similar metrics)
From paperRaw artifact
Field or section images captured from matched samples
Processed artifact
Selected representative panels with quantified intensity, counts, or area measurements
Final reported form
Per-group imaging summaries with representative figures and quantified endpoints
Hippocampal LTP magnitude and induction
From paperRaw artifact
Field or section images captured from matched samples
Processed artifact
Selected representative panels with quantified intensity, counts, or area measurements
Final reported form
Per-group imaging summaries with representative figures and quantified endpoints
Extracellular D-AP5 concentration in hippocampus via microdialysis
From paperRaw artifact
Field or section images captured from matched samples
Processed artifact
Selected representative panels with quantified intensity, counts, or area measurements
Final reported form
Per-group imaging summaries with representative figures and quantified endpoints
Total tissue D-AP5 concentration
From paperRaw artifact
Field or section images captured from matched samples
Processed artifact
Selected representative panels with quantified intensity, counts, or area measurements
Final reported form
Per-group imaging summaries with representative figures and quantified endpoints
Acquisition
Capture matched images from the relevant tissue region using the same acquisition settings across samples.
Preprocessing / cleaning
Correlation analysis was performed between D-AP5-induced impairment of spatial learning and corresponding impairment of hippocampal LTP in vivo.
Scoring or quantification
Quantify the primary readouts for this experiment: Spatial learning performance in Morris Water Maze (escape latency, path length, or similar metrics); Hippocampal LTP magnitude and induction; Extracellular D-AP5 concentration in hippocampus via microdialysis; Total tissue D-AP5 concentration.
Normalization
Normalize image-derived measurements against the matched acquisition or segmentation rules before comparing groups.
Statistical comparison
Statistical method not yet structured for this page.
Reporting output
Report representative outputs alongside summary comparisons for Spatial learning performance in Morris Water Maze (escape latency, path length, or similar metrics), Hippocampal LTP magnitude and induction, Extracellular D-AP5 concentration in hippocampus via microdialysis, Total tissue D-AP5 concentration.
Source links and direct wording from the methods section for validation and deeper review.
Citation
S Davis et al. (1992). The NMDA receptor antagonist D-2-amino-5-phosphonopentanoate (D-AP5) impairs spatial learning and LTP in vivo at intracerebral concentrations comparable to those that block LTP in vitro. Journal of Neuroscience
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