Lawrence M. Grover

REM sleep deprivation inhibits LTP in vivo in area CA1 of rat hippocampus

Rapid eye movement (REM) sleep deprivation has previously been shown to interfere with normal learning and memory and to inhibit long-term potentiation (LTP) in vitro. Previous studies on REM sleep deprivation and LTP have relied on in vitro analysis in isolated brain slices taken from animals following several days of sleep deprivation. LTP in the hippocampus in situ may differ from LTP in vitro due to modulatory inputs from other brain regions, which are altered after REM sleep deprivation. Here, we examined LTP in unanesthetized, behaving animals on the first and second recovery days following REM sleep deprivation to determine if similar effects are seen in vivo as previously reported in vitro. We found that LTP was significantly impaired in REM sleep-deprived animals on the second recovery day but not the first recovery day. Our results extend previous findings by showing that REM sleep deprivation continues to affect hippocampal function for more than 24h following the end of deprivation. Our results also suggest the presence of a modulatory process not present in vitro. Our findings are not explained by stress during REM sleep deprivation because equivalent circulating corticosterone levels (an index of stress) were found during both REM sleep deprivation and control treatment.

LTP in hippocampal area CA1 is induced by burst stimulation over a broad frequency range centered around delta

Long-term potentiation (LTP) is typically studied using either continuous high-frequency stimulation or theta burst stimulation. Previous studies emphasized the physiological relevance of theta frequency; however, synchronized hippocampal activity occurs over a broader frequency range. We therefore tested burst stimulation at intervals from 100 msec to 20 sec (10 Hz to 0.05 Hz). LTP at Schaffer collateral-CA1 synapses was obtained at intervals from 100 msec to 5 sec, with maximal LTP at 350-500 msec (2-3 Hz, delta frequency). In addition, a short-duration potentiation was present over the entire range of burst intervals. We found that N-methyl-d-aspartic acid (NMDA) receptors were more important for LTP induction by burst stimulation, but L-type calcium channels were more important for LTP induction by continuous high-frequency stimulation. NMDA receptors were even more critical for short-duration potentiation than they were for LTP. We also compared repeated burst stimulation with a single primed burst. In contrast to results from repeated burst stimulation, primed burst potentiation was greater when a 200-msec interval (theta frequency) was used, and a 500-msec interval was ineffective. Whole-cell recordings of postsynaptic membrane potential during burst stimulation revealed two factors that may determine the interval dependence of LTP. First, excitatory postsynaptic potentials facilitated across bursts at 500-msec intervals but not 200-msec or 1-sec intervals. Second, synaptic inhibition was suppressed by burst stimulation at intervals between 200 msec and 1 sec. Our data show that CA1 synapses are more broadly tuned for potentiation than previously appreciated.

Metabotropic glutamate receptor antagonist, (R,S)-α-methyl-4-carboxyphenyglycine, blocks two distinct forms of long-term potentiation in area CA1 of rat hippocampus

The necessity of metabotropic glutamate receptors (mGluRs) in the induction of long-term potentiation (LTP) has recently been questioned. We examined the effect of (R,S)-alpha-methyl-4-caboxyphenylglycine (MCPG), a selective mGluR antagonist, on two independent forms of LTP. One form induced by a 25 Hz/1 s tetanus is solely N-methyl-D-aspartate (NMDA) receptor-dependent. The other form induced by four 200 Hz/0.5 s bursts in the presence of APV is NMDA receptor-independent. In both paradigms the presence of MCPG prevented the induction of LTP by afferent activation.

Effects of extracellular potassium concentration and postsynaptic membrane potential on calcium-induced potentiation in area CA1 of rat hippocampus

Long-lasting potentiation can be induced in area CA1 of hippocampus by a relatively brief (7-10 min) exposure to a higher (4.0 mM) than normal (2.0 mM) extracellular calcium concentration. We have found that long-lasting calcium-induced potentiation is dependent on extracellular potassium concentration. Slices exposed to high extracellular calcium in the presence of normal extracellular potassium (3.35 mM) showed a transient facilitation. Long-lasting potentiation was induced by exposure to high calcium only in slices also exposed to higher than normal extracellular potassium (6.25 mM). In intracellular experiments we found that injection of depolarizing current into postsynaptic neurons could substitute for high extracellular potassium. These results suggest that calcium-induced potentiation involves a postsynaptic, voltage-dependent mechanism. A similar conclusion has been reached for tetanus-induced potentiation. We also found that calcium-induced potentiation, like tetanus-induced potentiation, is not accompanied by an increase in postsynaptic input resistance.

Differential effects of NMDA receptor antagonist APV on tetanic stimulation induced and calcium induced potentiation

Enduring synaptic potentiation can be induced in area CA1 of hippocampus by tetanic stimulation and by exposure to a medium containing high Ca2+ concentration. Both tetanic stimulation and high Ca2+ induce potentiation through voltage-dependent, post-synaptic mechanisms. Tetanus-induced long-term potentiation (LTP) was blocked by 50 microM D,L-2-amino-5-phosphonovalerate (APV) as previously reported by others. In contrast, Ca2(+)-induced long-lasting potentiation was not reduced by 50 microM APV. Thus the mechanisms by which tetanic stimulation and exposure to high Ca2+ induce synaptic potentiation may differ.

Growth hormone rescues hippocampal synaptic function after sleep deprivation

Sleep is required for, and sleep loss impairs, normal hippocampal synaptic N-methyl-D-aspartate (NMDA) glutamate receptor function and expression, hippocampal NMDA receptor-dependent synaptic plasticity, and hippocampal-dependent memory function. Although sleep is essential, the signals linking sleep to hippocampal function are not known. One potential signal is growth hormone. Growth hormone is released during sleep, and its release is suppressed during sleep deprivation. If growth hormone links sleep to hippocampal function, then restoration of growth hormone during sleep deprivation should prevent adverse consequences of sleep loss. To test this hypothesis, we examined rat hippocampus for spontaneous excitatory synaptic currents in CA1 pyramidal neurons, long-term potentiation in area CA1, and NMDA receptor subunit proteins in synaptic membranes. Three days of sleep deprivation caused a significant reduction in NMDA receptor-mediated synaptic currents compared with control treatments. When rats were injected with growth hormone once per day during sleep deprivation, the loss of NMDA receptor-mediated synaptic currents was prevented. Growth hormone injections also prevented the impairment of long-term potentiation that normally follows sleep deprivation. In addition, sleep deprivation led to a selective loss of NMDA receptor 2B (NR2B) from hippocampal synaptic membranes, but normal NR2B expression was restored by growth hormone injection. Our results identify growth hormone as a critical mediator linking sleep to normal synaptic function of the hippocampus.

Role of adenosine in heterosynaptic, posttetanic depression in area CA1 of hippocampus

Conditioning stimulation of afferent fibers in hippocampal area CA1 produced heterosynaptic, posttetanic depression (PTD) of responses evoked by stimulation of an independent set of afferent fibers. PTD was present within 5 s of conditioning stimulation, amounted to a 60-80% reduction of excitatory postsynaptic potentials (EPSPs), and required a period of 3-5 min for recovery. Antagonists of A1 adenosine receptors substantially reduced PTD. Adenosine released into, or formed in, the extracellular space during conditioning stimulation may diffuse within the slice to depress evoked release of glutamate.

Decreased afferent excitability contributes to synaptic depression during high-frequency stimulation in hippocampal area CA1.

Long-term potentiation (LTP) is often induced experimentally by continuous high-frequency afferent stimulation (HFS), typically at 100 Hz for 1 s. Induction of LTP requires postsynaptic depolarization and voltage-dependent calcium influx. Induction is more effective if the same number of stimuli are given as a series of short bursts rather than as continuous HFS, in part because excitatory postsynaptic potentials (EPSPs) become strongly depressed during HFS, reducing postsynaptic depolarization. In this study, we examined mechanisms of EPSP depression during HFS in area CA1 of rat hippocampal brain slices. We tested for presynaptic terminal vesicle depletion by examining minimal stimulation-evoked excitatory postsynaptic currents (EPSCs) during 100-Hz HFS. While transmission failures increased, consistent with vesicle depletion, EPSC latencies also increased during HFS, suggesting a decrease in afferent excitability. Extracellular recordings of Schaffer collateral fiber volleys confirmed a decrease in afferent excitability, with decreased fiber volley amplitudes and increased latencies during HFS. To determine the mechanism responsible for fiber volley changes, we recorded antidromic action potentials in single CA3 pyramidal neurons evoked by stimulating Schaffer collateral axons. During HFS, individual action potentials decreased in amplitude and increased in latency, and these changes were accompanied by a large increase in the probability of action potential failure. Time derivative and phase-plane analyses indicated decreases in both axon initial segment and somato-dendritic components of CA3 neuron action potentials. Our results indicate that decreased presynaptic axon excitability contributes to depression of excitatory synaptic transmission during HFS at synapses between Schaffer collaterals and CA1 pyramidal neurons.

Fitting experimental data to models that use morphological data from public databases

Ideally detailed neuron models should make use of morphological and electrophysiological data from the same cell. However, this rarely happens. Typically a modeler will choose a cell morphology from a public database, assign standard values for Ra, Cm, and other parameters and then do the modeling study. The assumption is that the model will produce results representative of what might be obtained experimentally. To test this assumption we developed models of CA1 hippocampal pyramidal neurons using 4 different morphologies obtained from 3 public databases. The multiple run fitter in NEURON was used to fit parameter values in each of the 4 morphological models to match experimental data recorded from 19 CA1 pyramidal cells. Fits with fixed standard parameter values produced results that were generally not representative of our experimental data. However, when parameter values were allowed to vary, excellent fits were obtained in almost all cases, but the fitted parameter values were very different among the 4 reconstructions and did not match standard values. The differences in fitted values can be explained by very different diameters, total lengths, membrane areas and volumes among the reconstructed cells, reflecting either cell heterogeneity or issues with the reconstruction data. The fitted values compensated for these differences to make the database cells and experimental cells more similar electrotonically. We conclude that models using fully reconstructed morphologies need to be calibrated with experimental data (even when morphological and electrophysiological data come from the same cell), model results should be generated with multiple reconstructions, morphological and experimental cells should come from the same strain of animal at the same age, and blind use of standard parameter values in models that use reconstruction data may not produce representative experimental results.

The modulation of excitatory synaptic transmission by adenosine in area CA1 of the rat hippocampus is temperature dependent

We tested the possibility that extracellular adenosine concentration varies with tissue temperature by measuring the tonic adenosinergic inhibition of excitatory synaptic transmission at different temperatures in the in vitro rat hippocampus. Application of the A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) enhanced population excitatory postsynaptic potentials (EPSPs) by antagonizing tonic adenosinergic inhibition; this effect was greatest at 25 degrees C, and was progressively reduced at 35 and 37.5 degrees C. These results demonstrate that tonic adenosinergic inhibition is inversely related to temperature. In a second experiment, an exogenous A1 agonist, N6-cyclohexyladenosine (CHA), was applied to slices to inhibit evoked EPSPs. CHA inhibition of EPSPs was greater at 35 than at 25 degrees C, demonstrating that the reduced adenosinergic inhibition at higher temperatures is not a result of reduced A1 receptor function.