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Keywords:

  • Epilepsy;
  • Animal Model;
  • Progesterone;
  • 3α,5α-THP

Abstract

  1. Top of page
  2. Abstract
  3. METHODS
  4. RESULTS
  5. DISCUSSION
  6. Acknowledgments
  7. REFERENCES

Summary: Purpose: Progestins can have profound effects on seizure processes. However, the effects and mechanisms of progestins to modulate seizures have not been systematically investigated. The present studies were designed to characterize the effects of progestins to modulate pentylenetetrazole (PTZ)-induced seizures in female rats.

Methods: In Experiment 1, ictal activity and plasma and hippocampal progesterone (P) and 5α-pregnan-3α-ol-20-one (3α,5α-THP) levels of proestrous rats were compared with those of diestrous and ovariectomized (ovx) rats. Experiments 2 and 3 examined effects of ovx and replacement with vehicle, P, or 3α,5α-THP, systemically (Experiment 2) or to the hippocampus (Experiment 3) on seizures and plasma and hippocampal P and 3α,5α-THP concentrations.

Results: Proestrous rats had reduced ictal activity and increased levels of P and 3α,5α-THP in plasma and hippocampus compared with diestrous or ovx rats (Experiment 1). Rats administered systemic P or 3α,5α-THP had significantly reduced ictal activity and increased plasma and hippocampal P and 3α,5α-THP levels compared with vehicle-administered rats (Experiment 2). Administration of P or 3α,5α-THP to the hippocampus of ovx rats significantly reduced seizure activity and increased hippocampal, but not plasma, levels of P and 3α,5α-THP compared with vehicle administration (Experiment 3).

Conclusions: Together, these data suggest that P can have antiseizure effects, and these effects may be due in part to actions of its metabolite, 3α,5α-THP, in the hippocampus.

Progesterone (P) and its 5α-reduced metabolite, 5α-pregnan-3α-ol-20-one (3α,5α-THP), have demonstrated antiseizure effects in some, but not all, reports (1). First, endogenous variations exist in seizure susceptibility of women and rodents, such that seizures are typically decreased during phases of the cycle when progestin levels are high compared with when they are low (2–7). Second, P or 3α,5α-THP administration to women with epilepsy, or to female rodents in seizure models, generally decreases ictal activity (8–16). Third, inhibiting the formation of 3α,5α-THP, by coadministering finasteride, a 5α-reductase inhibitor, with P increases seizures in people and animal models (3,10,17–19). Thus progestins may modulate seizures of women and rodents.

The antiseizure effects of progestins may be related to the formation of 3α,5α-THP in brain areas such as the hippocampus. The hippocampus is a target of progestins' actions. For example, P can protect the hippocampus from adrenalectomy (ADX)-induced neurodegeneration (20) and enhance neuroplasticity in the hippocampus (21). As well, 3α,5α-THP levels are highest in the hippocampus, compared with other brain areas examined (22). Although many brain areas are involved in the modulation of seizure processes, the hippocampus may be particularly sensitive to seizures. Seizures can result in degeneration of the hippocampus. Hippocampal neuron loss has been reported among children with severe seizures (23). Hippocampal damage also is reported in pilocarpine-, kainate-, pentylenetetrazole (PTZ)-, perforant pathway stimulation–, and kindling-induced ictal activity (14,24–26). Some evidence suggests that P or 3α,5α-THP administration can abrogate the effects of perforant pathway stimulation–induced seizures on neuron loss in the hilar region (10,24), although this is not the case in all reports (14). These data suggest that actions of progestins in the hippocampus may play a role in modulating ictal activity.

These data support a role of progestins in mediating seizures; however, whether actions of 3α,5α-THP in the hippocampus are involved in the antiseizure effects of progestins has not been directly investigated. Thus the current studies were designed to investigate the effects of progestins in the hippocampus to mediate PTZ-induced seizures in female rats. We hypothesized that if the antiseizure effects of P occur in part through actions of 3α,5α-THP in the hippocampus, then endogenous hormonal milieus or exogenous progestin regimens that increase 3α,5α-THP levels in the hippocampus would reduce PTZ-induced seizure activity.

METHODS

  1. Top of page
  2. Abstract
  3. METHODS
  4. RESULTS
  5. DISCUSSION
  6. Acknowledgments
  7. REFERENCES

These methods were preapproved by the Institutional Animal Care and Use Committee at SUNY-Albany.

Animals and housing

Female Long-Evans rats (N = 142), age ∼55 days, were obtained from the breeding colony at SUNY-Albany. Rats in Experiment 1 (n = 30) were left intact (proestrous and diestrous groups) or ovariectomized (ovx) at least 7 days before testing (ovx control group) and were group housed (four per cage). All rats for Experiments 2 (n = 30) and 3 (n = 30) were ovx at least 7 days before testing and group housed (four per cage). All rats were housed in a temperature-controlled room (21 ± 1°C) in the Laboratory Animal Care Facility. Rats were maintained on a 12/12-h reversed light cycle (lights off, 8 a.m.) with ad libitum access to Purina Rat Chow and tap water in their home cages. In addition to rats that were behaviorally tested, other rats that were in analogous conditions to those described earlier for each experiment were used to measure plasma and hippocampal P and 3α,5α-THP levels (n = 46; three to nine per group).

Determination of behavioral estrus

In Experiment 1, vaginal epithelium was obtained daily by lavage from intact rats ∼1 h before lights off and examined immediately by using low-power light microscopy. Rats were cycled through at least two normal estrous cycles (4- to 5-day cycle). Rats that had vaginal epithelium characterized by large round nucleated cells that are associated with proestrus (27) were subsequently placed in an arena with a sexually vigorous male that was allowed to mount the female 3 times to determine lordosis responsiveness (28). Rats were considered in proestrus (n = 10) if they had nucleated vaginal epithelium and demonstrated lordosis responses on at least one of three mount attempts. Rats that had vaginal epithelium containing neither nucleated nor cornified cells, that responded to attempted mounts by a stimulus male uniformly with lordosis ratings of 0, and that had not been proestrous the 2 previous days were considered in diestrus (n = 10). A control group of ovx rats was used in Experiment 1 (n = 10). Rats in Experiment 1 were tested within 4 h of lights out.

Surgery and hormone regimens

In Experiment 2, to control endogenous levels of hormones, rats were anesthetized with xylazine (Rompun; 12 mg/kg) and ketamine (Ketaset; 80 mg/kg) and ovx. One week after ovx, rats were given steroid hormones obtained from Sigma (St. Louis, MO, U.S.A.) and dissolved in sesame oil to yield a subcutaneous (s.c.) injection volume of 0.2 ml. Rats were administered P (500 μg, n = 10), 3α,5α-THP (500 μg, n = 10), or vehicle (sesame oil; n = 10) and tested 3 h later.

In Experiment 3, rats were ovx and stereotaxically implanted with bilateral guide cannulae aimed at the dorsal hippocampus (from bregma AP =–3.8; ML =± 2.0; DV =–2.0) (29). Cannula assembly consisted of 23-gauge thin-wall guide cannulae and 30-gauge removable inserts. One week after surgery, control inserts that were either tamped in cholesterol (or were left empty; n = 10), P (n = 10), or 3α,5α-THP (n = 10) were applied to the dorsal hippocampus. Our pilot research and previously published reports by others suggest that the diffusion of hormones from implants of this nature is ∼2 mm (unpublished data; 30). Immediately before application, inserts for experimental rats were verified as containing a 1-μg implant of steroid by checking the mass of the filled insert or confirming, with a dissecting microscope, or both, that the end of the insert was completely packed and no steroid was visible on the outside of the insert. Rats were tested 3 h after application of inserts.

PTZ model

PTZ is a tetrazol derivative with consistent convulsive actions in several animal models (31). The seizure progression produced with the PTZ regimen that we use (70 mg/kg, i.p.) is reliably observed and progresses in a manner similar to that described by Racine (32) for kindling-induced seizures. PTZ initially produces myoclonic jerks (mouth and facial twitches), followed by facial and forelimb clonus, which then becomes sustained and typically leads to generalized tonic–clonic seizures (loss of righting reflex and tonic forelimb flexion/extension followed by whole-body clonus) (33).

In these studies, the effects of progestins on PTZ-induced generalized seizure processes were examined. Two classes of motor convulsions are observed in experimental models of generalized seizures. Facial and forelimb clonic seizures, with or without rearing and falling, are often referred to as “forebrain” seizures. Alternatively, running–bouncing episodes followed by clonic–tonic convulsions that culminate in whole-body clonus with complete hindlimb extension require brainstem components at initiation (34). Forebrain and brainstem seizure circuits can maintain ictal activity independent of connections with each other; however, activation of either circuit can modify the other (35,36). Thus these studies examined whether progestins' actions in the hippocampus are sufficient to decrease generalized seizure processes of female rats.

PTZ is a useful model of generalized seizure disorder; however, individual variability can occur in the response to PTZ, such that ∼5% of rats are resistant to PTZ-induced seizures. The data of six rats in these experiments that did not show any evidence of myoclonus, facial and forelimb clonus, or tonic–clonic seizures after PTZ administration were excluded from data analyses (Experiment 1: n = 2 diestrus; Experiment 2: n = 1 ovx; Experiment 3: n = 2 P administered and n = 1 ovx). NB: Rats were not prescreened for seizure susceptibility, as it was previously demonstrated that ictal activity can alter 5α-reductase activity in the hippocampus and thereby neurosteroidogenesis, which could produce confounds in the investigation of the effects of progestins on ictal activity (37). To minimize variability in the effects of PTZ, we use the following procedures: The concentration of PTZ used (70 mg/ml) is made immediately before each test session to reduce possible degradation by light. Rats are carefully weighed immediately before testing to ensure that they receive precise dosing with PTZ (70 mg/kg). Controls are run in each test session to monitor that the effects of PTZ are consistent across test sessions. Uniform animal handling is practiced to obviate any possible stress-induced changes in seizures. Using these techniques, we have found that the antiseizure effects of progestins in the present report are similar to effects seen in other seizure models we have used in our laboratory (i.e., kainic acid or perforant-pathway stimulation or both) (12,24).

Behavioral testing

Immediately before testing, rats were weighed to ensure accurate dosing of PTZ. Rats were then placed in a plastic arena (50 × 30 × 25 cm) for 5 min to habituate. After habituation, rats were administered PTZ (70 mg/kg, i.p.). Immediately after PTZ injection, ictal behaviors were recorded for 10 min. Typically, within seconds of PTZ administration, myoclonus, characterized by mouth and facial movements and head nodding, was initiated. This was followed by facial and forelimb clonus and then tonic–clonic seizures, which were characterized by loss of righting reflex and tonic forelimb flexion/extension followed by whole-body clonus. In these studies, no consistent effects of progestins were noted on myoclonus or facial and forelimb clonus. However, the latency to, and the number of, tonic–clonic seizures was robustly altered by progestins. Thus ictal activity reported in these experiments includes the latency to, and number of, tonic–clonic seizures.

Tissue collection for hormone measurement

Tissues of rats in analogous conditions to those described earlier for each experiment were collected in the same time frame after progestin administration as for those rats assessed for ictal activity. This allowed plasma and hippocampal P and 3α,5α-THP levels produced by these regimens to be determined. Rats in each condition were rapidly decapitated, and trunk blood was collected and remained on ice until refrigerated centrifugation (4°C at 3,000 g for 8 min). Serum was aliquoted and stored at –70° C until radioimmunoassay for progesterone and 3α,5α-THP. Brains were rapidly removed; the hippocampus was dissected bilaterally, placed in dry ice, and stored at –70°C until radioimmunoassay for progesterone and 3α,5α-THP.

Radioimmunoassay for hormone measurement

Plasma and hippocampal P were determined by radioimmunoassay according to previously published methods (22). In brief, P was extracted from plasma samples with diethyl ether; the solvent was removed by using a speed drier, and samples were resuspended in assay buffer (pH 7.4). For brains, hippocampal tissue was homogenized with a glass/Teflon homogenizer in distilled water. Steroids were extracted from the homogenate with 50% MeOH, 1% acetic acid, dried down in an evaporator drier, and the pellet was reconstituted in trimethyl pentane (TMP) to half the homogenate volume. P was extracted from the reconstituted plasma and brain extracts by using Celite column chromatography. The progestin fraction was collected by using a 100% TMP wash. Fractions were dried by using a speed drier and then reconstituted in phosphate assay buffer. Radioimmunoassay was performed by using [3H]P (NET-208; specific activity, 48.4 Ci/mmol; New England Nuclear, Boston, MA, U.S.A.) and antisera (P#337 from Dr. G.D. Niswender, Colorado State University). The P antibody was used in a 1:30,000 dilution and bound between 30% and 50% of [3H]P. The standard curve was prepared in duplicate to give a range of nine concentrations from 50 to 8,000 pg/ml; total volume, 800 μl. Incubation (4°C for 24 h) was terminated by the addition of charcoal. After a 15-min incubation on ice, samples were centrifuged at 1,200 g for 10 min. Sample concentrations were calculated by using the logit–log method (38). The minimum detectable limit of the assay was 50 pg, and the intraassay and interassay coefficients of variance were 10.7% and 9.2%, respectively.

3α,5α-THP was measured according to previously established methods (22,39). In brief, steroids were extracted from plasma samples by using diethyl ether. Steroids were extracted from homogenized brain samples in 50% MeOH, 1% acetic acid, through a series of centrifugation and filtrations. Three hundred microliters of 0.1 M phosphate assay buffer (pH 7.4) was added to test tubes containing steroid extracts and equilibrated. The antibody, purchased from Dr. Robert Purdy (Veterans Medical Affairs, La Jolla, CA, U.S.A.), is specific to 3α,5α-THP (39). Although cross-reactivity of this 3α,5α-THP antibody occurs with 3α-hydroxypregn-4en-20-one (84%), dihydroprogesterone (11%), its β isomer (7%), and P (6%), negligible cross-reactivity (<2%) occurs with pregnenolone and corticosterone, as well as other progestins. Extensive cross-reactivity also is seen with the 5α-reduced metabolites of corticosterone and deoxycorticosterone. However, this antibody has been deemed suitable for measurement of 3α,5α-THP without high-performance liquid chromatography purification (39). The 1:5,000 dilution of this antibody bound between 40 and 60% of [3H]3α,5α-THP (NET-1047, 51.3 Ci/mmol; NEN). The standard curve was prepared in duplicate with a range of nine concentrations from 50 to 8,000 pg/ml; total volume, 950 μl. Incubation at 4°C for 24 h was terminated by charcoal separation of bound and free. Sample tube concentrations were calculated by using the logit–log method of Rodbard and Hutt (38), interpolation of the standards, and correction for recovery. The minimum detectable limit of the assay was 100 pg. The intraassay and interassay coefficients of variance were 0.12 and 0.15, respectively.

Statistical analyses

Effects of hormonal milieu on the latency to, and incidence of, seizures and plasma and hippocampal progestin levels were compared across groups in each experiment by using one-way analyses of variance (ANOVA). Alpha level for statistical significance was p ≤ 0.05. Where appropriate, Fisher's post hoc tests were used to determine group differences.

RESULTS

  1. Top of page
  2. Abstract
  3. METHODS
  4. RESULTS
  5. DISCUSSION
  6. Acknowledgments
  7. REFERENCES

Experiment 1

The latency to the first tonic–clonic seizure varied among groups with different endogenous progestin levels [F(2, 27) = 29.01; p < 0.01]. Rats in proestrus had significantly longer latencies to the first tonic–clonic seizure than did ovx rats and tended to have longer latencies than diestrous rats (see Fig. 1, top left).

image

Figure 1. Top: Latency to the first tonic–clonic seizure (left) and the mean number of tonic–clonic seizures (right) of ovx control (white bar), diestrous (striped bar), or proestrous (black bar) rats. *Significantly different from ovx control (p < 0.05). #Tendency for difference from diestrous (p < 0.10). Data are exclusive of two diestrous rats that did not exhibit any myoclonic, clonic, or tonic–clonic activity after pentylenetetrazol administration. Bottom: Hippocampal levels of progesterone (P; left) and 5α-pregnan-3α-ol-20-one (3α,5α-THP; right) of ovx control (white bar), diestrous (striped bar), or proestrous (black bar) rats. *Significantly different from ovx control (p < 0.05). #Significantly different from diestrous (p < 0.05).

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The number of tonic–clonic seizures varied as a function of hormonal milieu [F(2, 27) = 7.10; p < 0.01]. Proestrous rats had significantly fewer tonic–clonic seizures than did ovx rats and tended to have fewer seizures than diestrous rats (see Fig. 1, top right).

Proestrous rats had increased levels of plasma P [F(2, 13) = 23.7; p < 0.01] and 3α,5α-THP [F(2, 13) = 9.2; p < 0.01; data not shown] compared with diestrous or ovx rats. Proestrous rats also had increased P [F(2, 13) = 17.3; p < 0.01; see Fig. 1, bottom left] and 3α,5α-THP [F(2, 13) = 13.2; p < 0.01; see Fig. 1, bottom right] levels in the hippocampus compared with ovx or diestrous rats.

Experiment 2

The latency to the first tonic–clonic seizure varied across groups administered systemic progestins [F(2, 27) = 3.60; p < 0.05]. Rats administered SC P or 3α,5α-THP had significantly longer latencies to the first tonic–clonic seizure than did rats administered vehicle (see Fig. 2, top left).

image

Figure 2. Top: Latency to the first tonic–clonic seizure (left) and the mean number of tonic–clonic seizures (right) of ovariectomized (ovx) rats administered s.c. vehicle (ovx control; white bar), progesterone (P; striped bar), or 5α-pregnan-3α-ol-20-one (3α,5α-THP; black bar). *Significantly different from ovx control (p < 0.05). Data exclude one ovx control rat that did not exhibit any myoclonic, clonic, or tonic–clonic seizures after pentylenetetrazol administration. Bottom: Levels of hippocampal P (bottom, left) and 3α,5α-THP (bottom, right) of ovx rats administered s.c. vehicle (ovx control; white bar), P (striped bar), or 3α,5α-THP (black bar). *Significantly different from ovx control (p < 0.05). #Significantly different from 3α,5α-THP (p < 0.05).

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The incidence of tonic–clonic seizures varied as a function of systemic hormone administration [F(2, 27) = 7.13; p < 0.01]. Rats administered s.c. P or 3α,5α-THP had significantly fewer tonic–clonic seizures than did rats administered vehicle (see Fig. 2, top right).

Administration of s.c. progestins altered plasma P [F(2, 16) = 44.3; p < 0.01] and 3α,5α-THP [F(2, 16) = 5.2; p < 0.01; data not shown] levels compared with vehicle administration. The s.c. progestin administration also increased hippocampal P [F(2, 16) = 33.8; p < 0.01; see Fig. 2, bottom left] and 3α,5α-THP [F(2, 16) = 4.1; p < 0.01; see Fig. 2, bottom right) levels compared with those of vehicle. Notably, P administration increased P and 3α,5α-THP levels, whereas administration of 3α,5α-THP increased 3α,5α-THP, but not P, levels.

Experiment 3

The latency to the initial tonic–clonic seizure varied among groups administered progestins to the hippocampus [F(2, 27) = 29.01; p < 0.01]. Rats administered 3α,5α-THP to the hippocampus had significantly longer latencies to the first tonic–clonic seizure than did rats administered P or vehicle. A tendency was found for rats administered P to the hippocampus to have longer latencies than ovx rats (see Fig. 3, top left).

image

Figure 3. Top: Latency to the first tonic–clonic seizure (left) and the mean number of tonic–clonic seizures (right) of ovariectomized (ovx) rats administered vehicle (ovx control; white bar), progesterone (P; striped bar), or 5α-pregnan-3α-ol-20-one ((α,5α-THP; black bar) to the hippocampus. *Significant difference from ovx control (p < 0.05). #Tendency to be different from ovx control (p < 0.10). Data are exclusive of one ovx control rat and two P-administered rats that did not exhibit any myoclonic, clonic, or tonic–clonic seizures after pentylenetetrazol administration. Bottom: Hippocampal levels of P (bottom, left) and 3α,5α-THP (bottom, right) of ovx rats administered vehicle (ovx control; white bar), P (striped bar), or (α,5α-THP (black bar) to the hippocampus. *Significantly different from ovx control (p < 0.05). #Significantly different from 3α,5α-THP (p < 0.05).

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The incidence of tonic–clonic seizures was different among groups administered progestins to the hippocampus [F(2, 27) = 7.10; p < 0.01]. Administration of P or 3α,5α-THP to the hippocampus significantly decreased the number of tonic–clonic seizures compared to vehicle administration (see Fig. 3, top right).

Rats administered P or 3α,5α-THP to the hippocampus had significantly altered hippocampal P [F(2, 10) = 30.8; p < 0.01; see Fig. 3, bottom left] and 3α,5α-THP [F(2, 10) = 15.2; p < 0.01; see Fig. 3, bottom right], but not plasma (data not shown), progestin levels. Administration of P to the hippocampus increased P and 3α,5α-THP levels in the hippocampus. However, 3α,5α-THP administration to the hippocampus increased 3α,5α-THP, but not P, levels in the hippocampus.

DISCUSSION

  1. Top of page
  2. Abstract
  3. METHODS
  4. RESULTS
  5. DISCUSSION
  6. Acknowledgments
  7. REFERENCES

The present findings supported our hypothesis that actions of progestins in the hippocampus mediate the antiseizure effects of progestins in the PTZ seizure model. First, PTZ-induced ictal activity was decreased with endogenous hormonal milieus characterized by high levels of P and 3α,5α-THP in plasma and hippocampus. Second, systemic administration of a progestin regimen that produced high concentrations of P and 3α,5α-THP in plasma and hippocampus resulted in decreased seizure activity. Finally, intrahippocampal progestin administration had effects to decrease ictal activity similar to those seen after systemic administration but increased concentrations of P and 3α,5α-THP only in the hippocampus (not in plasma). Together, these data suggest that progestins can have antiseizure effects and that these effects may be due in part to actions of progestins in the hippocampus.

These findings confirm and extend past research that suggests that progestins can have antiseizure effects in various animal models of seizure disorder. Proestrous rats and ovx rats administered P or 3α,5α-THP had reduced ictal activity compared with relative controls. Notably, levels of 3α,5α-THP produced by our systemic and intrahippocampal replacement regimen produced 3α,5α-THP concentrations in the hippocampus well within physiologic ranges (less than that seen in proestrous rats) and decreased ictal activity (greater than that of proestrous rats). Previous reports demonstrated that progestins can decrease forebrain- as well as brainstem-mediated seizures; however, these studies used progestin regimens that produce pharmacologic progestin concentrations (40,41). The present data suggest that progestin regimens that produce physiologic concentrations of 3α,5α-THP in the hippocampus are sufficient to decrease PTZ-induced tonic–clonic seizures.

Although the endogenous and exogenous hormonal milieus that produced antiseizure effects in these studies were within the physiologic range, other endogenous factors also must be considered. For example, in Experiment 1, hippocampal progestin levels of ovx and diestrous rats were similar; however, ictal activity of diestrous rats was not as robustly increased as that of ovx rats, compared with proestrous rats. It is possible that the recent exposure (3–4 days previously) of diestrous rats to high levels of progestins may have had some protective effects, whereas ovx rats had been exposed to few or no progestins for 7 days. Thus recent prior exposure to progestins may reduce vulnerability to PTZ-induced seizures. As well, other hormones produced by the ovaries of intact rats, adrenals of ovx rats, or de novo synthesis in glial cells may account for some of the variability in the antiseizure effects of progestins that were seen in latencies of ovx groups in Experiment 2 compared with Experiments 1 and 3. Estrogen also is increased on proestrus and typically has proconvulsant effects [although not in all cases (i.e., 14,42)], which might explain the discrepancy between progestin levels and ictal activity seen in proestrous rats, compared with ovx, progestin-replaced rats, in the present studies. Further, among the ovx control rats, hippocampal levels of 3α,5α-THP were slightly higher than hippocampal P levels. Although it is possible that de novo production of 3α,5α-THP in the hippocampus may account for this, a more likely explanation is that a portion of the hormone measured in the hippocampal tissues of ovx rats represents the 5α-reduced metabolites of corticosterone. Rats in the present studies were not adrenalectomized, and the antibody used for radioimmunoassay cross-reacts with these metabolites. Notably, 5α-reduced metabolites of corticosterone have effects similar to those of 3α,5α-THP on γ-aminobutyric acid (GABA)ergic transmission (43). Thus although these data support a role of 3α,5α-THP in mediating seizures, other endogenous factors also may be important for modulating seizure processes.

Our current findings that P and 3α,5α-THP similarly reduce ictal activity suggest that some of the effects of P to modulate seizure processes may be due in part to actions of the 5α-reduced metabolite of P, 3α,5α-THP, in the hippocampus. Systemic or intrahippocampal administration of P or 3α,5α-THP similarly reduced ictal activity compared with vehicle-administered rats. Notably, whereas endogenous and systemic hormonal milieus produced higher plasma progestin levels, direct hippocampal progestin administration resulted in increased hippocampal, but not plasma, progestin levels. As well, direct implants of steroids to the hippocampus result in limited diffusion (∼2 mm) of the steroid, which would preclude progestins having actions in other brain regions to produce their antiseizure effects in Experiment 3. Further, when P or 3α,5α-THP was administered directly to the hippocampus, 3α,5α-THP appeared to have greater effects, albeit not significantly different, to reduce seizures compared with those of P. Notably, P has a high affinity for intracellular progestin receptors. However, in physiologic concentrations, 3α,5α-THP does not (44). 3α,5α-THP is a very potent modulator of GABAA receptors (45) and can alter the function of N-methyl-d-aspartate (NMDA) receptors (46). Thus the extent to which the antiseizure effects of progestins may be due in part to actions of 3α,5α-THP at intracellular progestin, GABAA and/or NMDA receptors in the hippocampus (or other brain areas) require further investigation.

In summary, these data support a role of progestins' actions in the hippocampus to modulate seizure processes are intriguing and have clear clinical relevance. In the United States, 2.5 million people have epilepsy; however, 30% of people with epilepsy do not respond to traditional pharmacotherapies used to control seizure activity. Further investigation of the effects and mechanisms of the ability of gonadal hormones to modulate ictal activity is necessary to expand the therapeutic options available to those with epilepsy.

Acknowledgments

  1. Top of page
  2. Abstract
  3. METHODS
  4. RESULTS
  5. DISCUSSION
  6. Acknowledgments
  7. REFERENCES

Acknowledgment:  This research was supported by a dissertation fellowship from The Epilepsy Foundation and the National Science Foundation (98-96263; 03-16083). The valuable input provided by dissertation committee members, Drs. Jacob Harney, Andrew Herzog, and Bruce Svare, is greatly appreciated.

REFERENCES

  1. Top of page
  2. Abstract
  3. METHODS
  4. RESULTS
  5. DISCUSSION
  6. Acknowledgments
  7. REFERENCES