Physiological and anatomical correlates of the human dentate gyrus: consequences or causes of epilepsy.

The presence of paroxysmal discharges in the epileptic human dentate gyrus provides a physiologic basis for hyperexcitability that may initiate seizure discharges during the development of epilepsy. Although these responses can occur with single orthodromic stimulation, data obtained under conditions that weaken synaptic inhibition (e.g., 1 Hz stimulation or bicuculline disinhibition) suggest that paroxysmal discharges may be a more common feature of tissue from temporal lobe epileptic patients than has been reported previously. Hilar cell loss and weakened synaptic inhibition may provide conditions favorable for the activation of N-methyl-D-aspartate acid (NMDA) receptors that would allow triggering of paroxysmal discharges that normally never are evoked in dentate granule cells in nonepileptic humans. As the dentate gyrus in normal animal tissue is not susceptible to intrinsic bursting behavior and is characterized by a relatively short duration excitatory postsynaptic potential even under pharmacologic disinhibition, paroxysmal discharges in the epileptic human dentate gyrus may provide an important clue to understanding the prerequisite conditions for seizure discharge.

Paroxysmal discharges in the EL mouse, a genetic model of epilepsy.

The EL/Suz (EL) mouse is a strain that is highly susceptible to convulsive seizures after repeated sensory stimulation. Its control strain, DDY/Jc1 (DDY), is less susceptible under similar conditions. The seizure prone phenotype is the result of differences at several genetic loci. In vivo electrical recordings from the seizure prone EL mouse brain have shown that the appearance of abnormal discharges in the hippocampus are critical to the onset of generalized seizures, indicating that the hippocampus plays an important role in EL mouse seizure activity. In the present study, electrophysiological differences between EL and DDY mice (9-15 weeks of age) were examined by comparing field potentials recorded from the dentate granule cell layer of hippocampal brain slices from mice that had not been stimulated to induce seizures. In control physiological solution, no significant differences were observed in characteristics of perforant path evoked field potentials or in paired pulse depression of evoked field potentials using 20 to 300 ms interstimulus intervals. After 60 min of disinhibition following bicuculline (10 microM) exposure, however, prolonged large amplitude potentials, paroxysmal discharges, were evoked by perforant path stimulation in the dentate gyrus of EL mice but were absent in the DDY strain. Paroxysmal discharges were curtailed by APV and were similar to responses recorded from the dentate gyrus in hippocampal brain slices from temporal lobe epileptic patients. The field response to hilar stimulation was identical in both strains and was composed of a single population spike before and after bicuculline exposure. Mossy fiber terminals were not present in the molecular layer of either strain. We propose that the mechanisms leading to a greater likelihood of paroxysmal discharge generation in EL mouse may be important in the development and/or generation of epileptic seizures in this mouse strain and may be a significant phenotypic difference between the EL mouse and its parent strain.

Mossy fiber reorganization and its possible physiological consequences in the dentate gyrus of epileptic humans.

It is unlikely that MF reorganization is the cause of epilepsy, but it may affect the progression of the disease, i.e., the frequency or severity of seizures. We propose that early events, yet undiscovered, lead to an increased likelihood of excitability. This hyperexcitability, which initially may not be manifested in overt seizures, may erode vulnerable hilar neurons that serve an important inhibitory function, as illustrated in Fig. 6-15. As inhibition is lost, hyperexcitability reaches the level of clinically manifested seizures that are severe enough to lead to substantial loss of hilar neurons. When the loss of these cells is sufficiently high, MF reorganization occurs, first to neighboring hilar neurons and later to dendrites of granule cells (Fig. 6-15). Thus, the functional consequence of MF reorganization may provide a compensatory form of inhibition, as well as a circuit for feedback excitation. Although definitive evidence indicating that MF reorganization contributes to the acceleration or progression of epilepsy is missing, the findings to date are consistent with this hypothesis. In the event that reorganization contributes to the epileptic condition, treatments that reduce indicators of neuropathology may lead to a reduction of seizure frequency and severity. Evidence suggests that reorganization in the dentate gyrus may follow the pathways of neuronal processes of hilar neurons that have died. Thus, further study of the events that guide MF reorganization may hold important clues for developing methods for targeting regenerating axons following central nervous system injury.

Enhancement of paired-pulse depression in the dentate gyrus in vivo by the NMDA antagonist, MK-801, and electrical kindling.

We have previously demonstrated that late paired-pulse depression of dentate granule cell field potentials decreases when stimulus intensity is increased from moderate to high levels. Voltage-dependent N-methyl-D-aspartate (NMDA) currents are increasingly activated within this stimulus range, and are enhanced following the development of kindled seizures. The NMDA antagonist, MK-801 (0.25 and 1.0 mg/kg, i.p.), was used in the present experiments to evaluate the contribution of NMDA currents to the loss of late paired-pulse depression at high stimulus intensities in naive and kindled rats. Paired-pulse stimulus intensity functions were obtained from animals prepared with chronic electrodes in the perforant path and dentate gyrus. MK-801 administration had no effect on the stimulus intensity function for early paired-pulse depression (20-30 ms interpulse intervals, IPI) in either preparation. Late paired-pulse depression (150-500 ms IPI) was significantly enhanced in naive rats by MK-801. In contrast, MK-801 had no effect on the potentiation of late paired-pulse depression recorded from kindled animals. These findings suggest that the ability of NMDA currents to reduce the strength of late paired-pulse depression in naive animals is altered following the development of kindled seizures. A decrease in late paired-pulse depression was observed at high stimulus intensities under all experimental conditions. The latter findings indicate that the processes responsible for the reduction in late paired-pulse depression at high stimulus intensities are unaffected by either NMDA or kindling-induced modulation of late paired-pulse depression.

Changes in dentate granule cell field potentials during afterdischarge initiation triggered by 5 Hz perforant path stimulation.

A failure of early paired pulse depression often precedes the onset of intermittent spontaneous seizures in animal models of status epilepticus. In the present study, changes in the strength of early and late paired pulse depression of dentate granule cell field potentials were compared in the unanesthetized rat during the initiation of a single afterdischarge (AD) evoked by perforant path stimulation (0.1 ms pulse duration, 5 Hz, 12-18 s duration, 50-1000 microA). Late paired pulse depression was measured by sequential changes in the population spike (PS) amplitude during 5 Hz stimulation (200 ms interpulse interpulse interval, IPI). When 5 Hz stimulation triggered an AD, the population spike (PS) was initially depressed and then increased to above pre-train values, indicating a loss of late paired pulse depression by the middle of the train. Early paired pulse depression was measured by inserting paired pulses (20 ms IPI) at spaced intervals throughout the 5 Hz train. In contrast to late paired pulse depression, early paired pulse depression remained at maximum strength until an abrupt failure was detected coincident with AD initiation. Two experimental treatments shown to increase the strength of late paired pulse depression, administration of the N-methyl-D-aspartate antagonist, MK-801 (0.25 mg/kg, i.p.), and the development of kindled seizures, produced an increase in AD thresholds and in the initial depression in the PS amplitude during 5 Hz stimulation. Together, these results suggest that a failure of late paired pulse depression may be a precipitating event in AD initiation triggered by 5 Hz stimulation in the unanesthetized rat.

Stimulus parameters affecting paired-pulse depression of dentate granule cell field potentials. I. Stimulus intensity.

Paired pulse stimulation of the perforant path provides a measure of inhibition of dentate granule cell field potentials that is reflected in the depression of the second (test) population spike (PS) relative to the first (conditioning) PS. The assumption that the strength of paired pulse depression is dependent upon the amplitude of the conditioning PS was investigated by increasing the stimulus intensity of both pulses (5-100% of maximum, Experiment 1), or by increasing the stimulus intensity of the conditioning pulse (5-100%) while maintaining a constant stimulus intensity of the test pulse (50%, Experiment 2). In both experiments, the threshold for early paired pulse depression (20 ms interpulse interval, IPI) was reached with moderate stimulation (30-40% of maximum). Above threshold, the test PS was depressed to a relatively constant amplitude in Experiment 1, in contrast to a nearly linear decrease observed in Experiment 2 with increasing strength of early paired pulse, relative to the conditioning reflects the lower stimulus intensity of the test pulse, relative to the conditioning pulse, in the second study, thereby allowing the increasing strength of early paired pulse depression to be detected more easily. The threshold for late paired pulse depression was reached near (20%, Experiment 1) or below (5%, Experiment 2) the PS threshold of dentate granule cells, and a paradoxical decrease in late paired pulse depression was detected with maximal stimulation in both studies. Together, these results suggest that early paired pulse depression exhibits a strong dependence upon the amplitude of the conditioning PS, whereas late paired pulse depression is marginally affected by the conditioning PS amplitude and is influenced by additional processes at both extremes of the stimulus intensity continuum.

Stimulus parameters affecting paired-pulse depression of dentate granule cell field potentials. II. Low-frequency stimulation.

Low frequency (1 Hz) stimulation of the perforant path produces a depression in the population spike (PS) of dentate granule cell field potentials and also may affect the strength of paired pulse depression. The effects of 1 Hz stimulation (30 s train) on paired pulse depression (20 and 200 ms interpulse intervals, IPI) were evaluated in the unanesthetized rat under two conditions: (i) when the stimulus intensity of both pulses was increased simultaneously (5-100%); and (ii) when the stimulus intensity of the first (conditioning) pulse was increased (5-100%), while the stimulus intensity of the second (test) pulse was held constant (50%). The test PS amplitude was predicted based upon either the conditioning PS amplitude at the end of the 1 Hz train or upon the additive effects of paired pulse depression and 1 Hz stimulation. These predicted values then were assessed for the best fit to observed values following 1 Hz trains. Under both stimulus conditions, the 1 Hz depression in the conditioning PS amplitude exhibited characteristics that were identical to late paired pulse depression recorded before the train. A decrease in the test PS amplitude also was observed following 1 Hz stimulation at the 20 and 200 ms IPIs. The best fit to observed values of the test PS at the end of 1 Hz trains was provided by estimates based upon the additive effects of 1 Hz stimulation and paired pulse depression. These results indicate that the strength of paired pulse depression in the unanesthetized rat is unchanged following 1 Hz stimulation, and further, that the 1 Hz depression in dentate granule cell field potentials most likely reflects the cumulative influence of late paired pulse depression.

Longitudinal variation in cell density and mossy fiber reorganization in the dentate gyrus from temporal lobe epileptic patients.

Variation in cell loss and mossy fiber reorganization was examined along the longitudinal axis of the dentate gyrus from temporal lobe epileptic (TLE) patients. Previous evidence has indicated that the anterior hippocampus is prone to seizure activity. We compared granule and hilar cell number in addition to Timm stain density of the molecular layer and hilus in more anterior and more posterior specimens of hippocampus obtained from patients surgically treated for intractable epilepsy by the removal of the anterior half of the hippocampus. Granule cells/mm in the more anterior specimen were less than or equal to those in the more posterior specimen locations in 77% of the patients, while there was no significant difference in hilar neuron density between the two blocks. These results demonstrate a significantly greater pathology in the granule cell layer in more anterior specimens and no difference in pathology for hilar neurons. Molecular layer Timm stain density was significantly greater in the more anterior specimen of 71% of the patients. The molecular layer Timm stain density ratio was inversely related to hilar cell density in more anterior specimens, whereas in more posterior specimens there was no significant relationship with hilar cell density. Our observations show that although differences exist among TLE patients for these neuroanatomic measures, pathology was greater in more anterior specimens. The latter result is consistent with the conclusion that seizure activity may originate in the anterior region of the hippocampus in a majority of patients.

The functional relationship between antidromically evoked field responses of the dentate gyrus and mossy fiber reorganization in temporal lobe epileptic patients.

Field recordings from the dentate granule cell layer of in vitro brain slices of temporal lobe epileptic patients were evoked by antidromic stimulation. Tissue from the same specimen was stained by the Timm-sulfide method to assess the pattern and degree of mossy fiber reorganization into the supragranular layer. A wide range of physiological responses and Timm staining patterns was present across patients. A significant correlation was observed between the abnormality of antidromic responses, reflected by multiple secondary population spikes, and the degree of Timm staining of the supragranular layer. This relationship lends support to the hypothesis that mossy fiber synapses located in the supragranular layer may have functional implications for granule cell excitability in human epileptic tissue.

Ca2+/calmodulin-dependent protein phosphorylation is not altered by amygdaloid kindling.

Kindling is a process in which episodic electrical stimulation permanently increases seizure susceptibility. One mechanism to account for a change in seizure susceptibility is some alteration in signal transduction, possibly at the level of second messenger systems. In this study, male Long-Evans rats were kindled in the amygdala, and Ca2+/calmodulin (Ca2+/CaM)-dependent protein phosphorylation was assessed at the site of the primary kindled focus using one- and two-dimensional gel electrophoresis. In vitro phosphorylation of membrane and cytosol fractions in the presence of absence of Ca2+/CaM did not differentiate kindled from nonkindled amygdaloid tissue. These results suggest that changes in Ca2+/CaM-dependent phosphorylation are not related to the mechanism(s) underlying the establishment of an amygdaloid kindled focus.

Convulsant properties of cyclotrimethylenetrinitramine (RDX): spontaneous audiogenic, and amygdaloid kindled seizure activity.

Dose-effect and time course relationships were determined for the effects of the explosive cyclotrimethylenetrinitramine (RDX) on seizure susceptibility. Male Long Evans rats treated with 0-60 mg/kg RDX po were monitored for spontaneous seizures during an 8-hr interval between dosing and audiogenic (AG) seizure testing. Blood samples for analyzing plasma RDX concentrations were obtained immediately thereafter. Spontaneous and AG seizures were observed at dosages as low as 10-12.5 mg/kg, with significant seizure incidence induced by dosages of 25.0 mg/kg (5.34 micrograms RDX/ml plasma) and 50.0 mg/kg (8.28 micrograms RDX/ml plasma), respectively. Spontaneous seizure incidence peaked at 2 hr for all RDX treatment groups, then decreased (12.5 and 25.0 mg/kg) or remained elevated (50.0 mg/kg) for the remaining 6 hr. In contrast, AG seizures (37.5 mg/kg) could be elicited only at 8 and 16 hr, despite significant elevation of plasma RDX concentrations at 2 and 4 hr. Because limbic system involvement was suggested by spontaneous seizure characteristics, the rate of amygdaloid kindling was measured following daily treatment with 6.0 mg/kg. This dosage significantly accelerated kindling development without inducing spontaneous seizures or producing an accumulation of RDX in plasma. These data provide preliminary evidence that limbic structures may participate in RDX-induced seizure susceptibility.

Differential effects of caffeine, picrotoxin, and pentylenetetrazol on hippocampal afterdischarge activity and wet dog shakes.

We identified changes in hippocampal afterdischarge activity that follow administration of subcon vulsant doses (one-half the convulsant dose) of analeptic agents with known pharmacological action. Long-Evans rats (N = 104) with chronic bipolar electrodes implanted in the dorsal hippocampus, were injected i.p. with saline, caffeine (75 mg/kg), picrotoxin (2 mg/kg), or pentylenetatrazol (20 mg/kg) in 1 ml/kg volume 15 min before testing. Body temperature was measured at the beginning of the session to determine if significant change was associated with any of the treatments. Beginning at 10 microA, current (2-s train of 50-Hz biphasic pulses) was applied to hippocampal electrodes and intensity was increased in 10-microA steps until the afterdischarge sequence was elicited. Afterdischarge threshold, wet dog shake frequency, and the duration of the primary afterdischarge, the postprimary depression, and the rebound afterdischarge were measured. Caffeine administration produced a dramatic prolongation of the rebound afterdischarge, without affecting the duration of the primary afterdischarge. All other afterdischarge variables were unchanged by the caffeine treatment. Because caffeine blocks adenosine receptors at physiologic concentrations, adenosine action is implicated in the termination of the second, but not the first, spike train. Picrotoxin and pentylenetetrazol had no influence on the EEG, despite evidence of slight (1 degrees C) hypothermia. A decrease in wet dog shake frequency, however, was associated with picrotoxin administration. As picrotoxin and pentylenetetrazol are known gamma-aminobutyric acid (GABA) antagonists, the results suggest that GABA is involved minimally, if at all, in the hippocampal afterdischarge sequence.

Sulfolane effects on audiogenic, pentylenetetrazol and afterdischarge seizure activity.

Sulfolane dosages that alter seizure susceptibility were determined using audiogenic (AG), pentylenetetrazol (PTZ) and hippocampal afterdischarge (AD) seizure models. The presence of AG seizures and potentiation of PTZ seizures were investigated in rats injected IP with 0, 200, 400 or 800 mg/kg; AD activity was assessed only at the highest and lowest dosages. The dose-dependent hypothermia associated with sulfolane treatment was controlled in Experiment I and a replication study (Experiment II) by testing under isothermic conditions. The effect of body temperature was measured directly in Experiment III by comparing AG seizure incidence and characteristics exhibited by hypothermic and normothermic animals. Audiogenic seizures were elicited in nearly half of the 800 mg/kg animals in both Experiments I and II. Sulfolane-induced hypothermia, maximal at 3 hours, partially protected against AG seizure characteristics. Potentiation of PTZ seizure severity (800 mg/kg) and duration (800 and 400 mg/kg) also were observed. None of the hippocampal AD parameters was affected significantly by sulfolane treatment. The similarity of the convulsants sulfolane and PTZ is discussed.

Long-term behavioral and electrophysiological changes associated with lead exposure at different stages of brain development in the rat.

The present investigation was conducted to assess the behavioral and electrophysiological impairments exhibited by adult male rats as a function of the developmental stage during which lead exposure occurred. Dams were given either a lead acetate (0.3%) or a control drinking solution during days 16-23 of gestation, days 1-8 or days 9-16 of nursing. The temporal and spatial activity patterns exhibited by gestationally exposed offspring in the open field between 42-45 days of age was distinguished from all other groups by the absence of a decrement in peripheral field activity across days and by increased exploration of the center field. Although open field activity proved sensitive to the timing of lead exposure, power spectral analyses of hippocampal and cortical EEG activity at 70-72 days of age revealed that lead selectively depressed 6-7 Hz energy in the hippocampus, independent of the developmental stage of exposure; cortical EEG and other hippocampal theta frequencies were unaffected. The differential sensitivity of open field activity and select hippocampal theta frequencies to the timing of lead administration suggests that the identification of toxic consequences depends on the function assessed and the developmental stage during which lead exposure occurred.

A comparison of saccade evoked potentials recorded during reading and tracking tasks.

The influence of visual processing demands on saccade-triggered evoked potentials was investigated at P3, P4 and Oz recording sites during reading and tracking tasks. To maximize the physical similarities between tasks, subjects tracked a series of lights that flashed in a stereotypic reading pattern behind a page of text; eye movements recorded during reading initiated the light sequence. In the first experiment, a significant decrease observed in the latency of the major positive peak recorded from Oz during tracking was attributed to the smaller amplitude of tracking, relative to reading, saccades. To confirm this interpretation, the experiment was repeated with modification to the light display. As anticipated, equating saccade amplitudes across tasks eliminated waveform differences in the second experiment. Although peak latencies and amplitudes were not influenced reliably by visual processing demands, tracking potentials exhibited a negative DC shift relative to reading waveforms that was significant at 174 msec at the Oz site. These data suggest that the saccade-triggered evoked potential components generally are insensitive to task differences within the visual modality when visual configuration and eye movement parameters are controlled.

Neurophysiological consequences of acute exposure to methylpyridines.

A series of neurophysiological tests was performed on Long-Evans hooded rats treated with either 2-, 3-, or 4-methylpyridine at dosages of 100 mg/kg, approximately one-half the ip LD50. The tests contained measures of sensory function (paired pulse flash evoked potentials, pattern reversal evoked potentials, and brainstem auditory evoked responses) and cerebral excitability (pentylenetetrazol seizures and hippocampal afterdischarges). In general, rats treated with 2- and 3-methylpyridine were more affected than those treated with 4-methylpyridine. The changes observed were in many ways similar to those seen following administration of depressant compounds: increased latency of evoked potentials and increased latency to PTZ seizures. Not all findings, however, were consistent with previously observed patterns of central nervous system depression.

Locus of control and vasomotor response to sensory processing.

Heart rate and forearm blood flow responses were measured during experimental tasks requiring sensory intake, sensory rejection, and a mixture of the two behaviors. Subjects were 29 college students who had been categorized using Rotter's locus of control scale. Significant increases in both cardiovascular indices were seen in all three tasks; the responses were smaller, however, during sensory intake. Internals showed a differential response of forearm blood flow. In contrast, externals showed a similar vasodiatation across all tasks. The findings indicate that some of the variability in cariovascular response to sensory processing may be explained by individual differences in personality charcteristics related to subjects' perferred level of involvement in differing sensory processing behaviors.