neurophysiology tutorial paper 3

• the paper (Tsien et al, 1996) found that adult mice with a CA1-restricted gene knockout of NMDAR1 lack NMDA receptor mediated synaptic currents and LTP in CA1 synapses. Spatial learning was impaired but non-spatial learning was unaffected. results suggested that activity dependent modifications of CA1 synapses, mediated by NMDA receptors, play an essential role in acquisition of spatial memories.
• it had been long hypothesised that memory storage in the mammalian brain involves modifications of the synaptic connections between neurons. Hebb (1949) introduced the concept of correlated activity, which is when the presynaptic and postsynaptic neurons are active simultaneously, they become strengthened. NMDA receptors implement this at a synaptic level as they are coincidence detectors
• NMDA receptors are coincidence detectors as they require both presynaptic and postsynaptic changes to become active. presynaotically, glutamate must be released. postsynaptically, the AMPA receptors (+ other channels) must depolarise the membrane to relieve the Mg 2+ block of the NMDA channel.
• Long term potentiation is used as un indication of an increase in synaptic efficacy. induction requires activation of NMDARs. NMDAR dependent LTP is elicited by giving a strong pattern of electrical stimulation ( 25-100 Hz train for approx 1 second) to the inputs. rapid and lasting increase in synaptic strength.
• We have produced a mouse strain in which the deletion of the NMDAR1 gene is restricted to the CA1 pyramidal cells of the hippocampus by using a new and general method that allows CA1-restricted gene knockout
• The mutant mice grow into adulthood without obvious abnormalities
• Adult mice lack NMDA receptor–mediated synaptic currents and long-term potentiation in the CA1 synapses and exhibit impaired spatial memory but unimpaired nonspatial learning
• Our results strongly suggest that activity-dependent modifications of CA1 synapses, mediated by NMDA receptors, play an essential role in the acquisition of spatial memories
• NMDAR


  1. methyl-D-aspartate receptor
• LTP


  Long-term potentiation
• NMDARs can implement the Hebb rule at the synaptic level, and they are thus considered the crucial synaptic elements for the induction of activity-dependent synaptic plasticity
• NMDARs act as coincidence detectors because they require both presynaptic activity (glutamate released by axonal terminals) and postsynaptic activity (depolarization that releases the Mg2+ block) as a condition for channel opening
• Active NMDAR channels allow calcium influx into the postsynaptic cell, which triggers a cascade of biochemical events resulting in synaptic change
• Conventional NMDAR-dependent LTP is elicited by giving a strong pattern of electrical stimulation (a 25–100 Hz train for ~1 s) to the inputs, which triggers a rapid and lasting increase in synaptic strength
• The hippocampus is the most intensely studied region for the importance of NMDARs in synaptic plasticity and memory
• Lesions of the hippocampus in humans and other mammals produce severe amnesia for certain memories
• Disruption of NMDARs in the hippocampus leads to blockade of synaptic plasticity and also to memory malfunction
• Application of NMDAR antagonists (such as 2-amino-5-phosphonopropionic acid [AP5]) completely blocks the induction of LTP in most hippocampal synapses
• Rats that received infusion of AP5 into the hippocampus were deficient in performing a spatial memory task (Morris et al, 1986). animals were required to form spatial relations between a water maze and visible objects in the surrounding environment.
• Mice with a deletion in the gene encoding the α subunit of calcium-calmodulin-dependent protein kinase II (αCaMKII) display impaired LTP in the CA1 region of the hippocampus and a deficit in spatial learning
• The Cre/loxP recombination system was used to restrict the deletion of the NMDAR1 gene to the pyramidal cells of the hippocampal CA1 region. the CA1-KO mice had no obvious pathologies, unlike the general knockout mice which died neonatally (Lie et al, 1994)
• The CA1-KO mice lack NMDAR-mediated postsynaptic currents and LTP in the CA1 region
• The CA1-KO mice show impaired spatial memory (measured in the hidden-platform version of the Morris water maze) but display good performance in nonspatial learning tasks
• In mutant slices (Figure 5A, right) looked normal when compared with control slices (data not shown)
• After addition of a solution that isolates NMDARs, the mutant slices showed clear NMDA EPSPs (n = 12; Figure 5A, right) as well as the control slices (data not shown)
• The input–output relations were indistinguishable between the CA1-KO and control slices (Figure 5B)
• The paradigm of paired pulse facilitation, which gives an indication of presynaptic function, showed no differences between the mutant and control slices (Figure 5C)
• The CA1-KO cells lacked the slow component of the excitatory postsynaptic current (EPSC) that is mediated by NMDARs

  
• The early component of the EPSC that is mediated by AMPA receptors was intact in the mutants (Figure 4A)
• Tetanic stimulation (100 Hz for 1 s) failed to induce LTP in the CA1-KO slices (Figure 6A)
• Short-term potentiation was not detectable in the mutant slices
• On average, the synaptic responses were unchanged in CA1-KO slices 45 min after the tetanus (100.4 ± 4.9%, n = 21; Figure 6B), whereas LTP was readily obtained in control slices (T29–1, 168.5 ± 4.9%, n = 12; fNR1, 156.9 ± 8.3%, n = 4; wt, 177.7 ± 6.2%, n = 5; Figure 6B)
• Tetanic stimulation (40 shocks at 100 Hz) was able to induce LTP in either of the two inputs (lateral perforant path and medial perforant path) in the dentate gyrus of the CA1-KO mice (Figure 6C)

  
• On average, the lateral perforant path showed a statistically significant increase in synaptic efficacy 45 min after the tetanus (154.6 ± 10.1%, n = 10, p < 0.01, t test; Figure 6D)
• The CA1-KO mice were deficient in learning the spatial memory task in the water maze when compared to control siblings
• The CA1-KO mice did not show any place preference for the target quadrant in any of the transfer tests (TT1, TT2, TT3)
• The control groups showed a marked preference for the target quadrant in TT2 and remained at this same level of quadrant preference in TT3
• The CA1-KO mutants had significantly fewer crossings of the correct location of the platform during TT3 compared to control mice
• The CA1-KO mutants showed a significant deficit on the "platform search", the average time searching in the exact location of the platform, compared to control mice
• CA1-KO mice


  • Deficient in spatial memory
  • Showed complete deficit in spatial memory in water maze apparatus