- Email: [email protected]
Low frequency electrical stimulation has time dependent improving effect on kindling-induced impairment in long-term potentiation in rats
Accepted Manuscript Research report Low frequency electrical stimulation has time dependent improving effect on kindling-induced impairment in long-term potentiation in rats Khadijeh Esmaeilpour, Vahid Sheibani, Mohammad Shabani, Javad MirnajafiZadeh PII: DOI: Reference:
S0006-8993(17)30196-8 http://dx.doi.org/10.1016/j.brainres.2017.05.007 BRES 45356
To appear in:
Brain Research
Received Date: Revised Date: Accepted Date:
24 December 2016 28 April 2017 8 May 2017
Please cite this article as: K. Esmaeilpour, V. Sheibani, M. Shabani, J. Mirnajafi-Zadeh, Low frequency electrical stimulation has time dependent improving effect on kindling-induced impairment in long-term potentiation in rats, Brain Research (2017), doi: http://dx.doi.org/10.1016/j.brainres.2017.05.007
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Low frequency electrical stimulation has time dependent improving effect on kindling-induced impairment in long-term potentiation in rats
Khadijeh Esmaeilpour1, Vahid Sheibani1, Mohammad Shabani1, Javad Mirnajafi-Zadeh1,2*
1- Neuroscience Research Center, Institute of Neuropharmacology, Kerman University of Medical Sciences, Kerman, Iran 2- Department of Physiology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
*Corresponding author: Javad Mirnajafi-Zadeh, Department of Physiology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, PO Box: 14115-331, Iran, Tel: +98-21 82883865, Fax: +98-21 82884825, E-mail: [email protected]
1
Abstract Application of low-frequency stimulation (LFS) can improve learning and memory in kindled animals (Ghafouri et al., 2016). Considering the important role of long-term potentiation (LTP) in learning and memory, in the present study the effectiveness of LFS on kindling-induced impairment in LTP induction was investigated in hippocampal CA1 area at different times post kindling stimulations. Animals were kindled via electrical stimulation of hippocampal CA1 area in a semi-rapid manner (12 stimulations per day). One group of animals received four trials of LFS at 30 s, 6 h, 24 h, and 30 h following the last kindling stimulation. Each LFS consisted of 4 packages at 5 min intervals; each package contained 200 monophasic square wave pulses of 0.1 ms duration at 1 Hz. The kindled, kindled+LFS and LFS groups were divided into four subgroups in which hippocampal slices were prepared at 48 h, 1 week, 2 weeks, and 1 month following the last kindling stimulation respectively. Extracellular evoked field excitatory postsynaptic potentials (fEPSPs) were recorded in the stratum radiatum of the CA1 area of the slice. Obtained results showed that LTP was not induced in kindled animals. However, application of LFS overcame the kindling-induced impairment in LTP generation in CA1 area of the hippocampus. This improving effect remained up to one week after the last kindling stimulation and extended to one month by increasing the number of applied LFS packages.
Keywords: Seizure; Low frequency stimulation; Long-term potentiation; Hippocampus
2
1. Introduction Temporal lobe epilepsy is one of the most prevalent neurological disorders (Schiller and Bankirer, 2007; Volcy, 2003). Impairment in learning and memory has been reported in patients with epilepsy (Carreño et al., 2008; Holmes, 1991). Kindling is an experimental animal model for temporal lobe epilepsy which can be induced by daily low-intensity electrical stimulation of limbic areas such as the hippocampus (Goddard et al., 1969; Racine, 1978). Similar to epileptic patients, kindled animals show impairment in spatial learning and memory which maybe a result of damage to neuronal function in the hippocampus (Cammisuli et al., 1997; Da Silva et al., 1986; Gilbert et al., 1996; Hannesson et al., 2001; Leung et al., 1990). Long-term potentiation (LTP) in the hippocampus is a form of synaptic plasticity which has been considered as the neuronal mechanism involved in learning and memory (Bliss and Lømo, 1973; Bliss and Collingridge, 1993; Nabavi et al., 2014; Pastalkova et al., 2006; Soderling and Derkach, 2000; Teyler and DiScenna, 1985). Kindling phenomenon is accompanied with synaptic potentiation and there are many similarities between kindling-induced potentiation and LTP (Krug et al., 1997; McEachern and Shaw, 1996). Accordingly, a possible mechanism of kindling epileptogenesis is long-lasting enhancement of excitatory synapses (Baudry, 1985; Douglas and Goddard, 1975; Maru et al., 1982; Racine et al., 1975; Sutula and Steward, 1986). Many studies have indicated that kindled seizures can lead to synaptic potentiation which is accompanied with an increase in slope of field excitatory postsynaptic potential (fEPSP) and amplitude of population spike (PS) (Jahanshahi et al., 2009; Mohammad-Zadeh et al., 2007; Mohammad-Zadeh et al., 2009; Robinson et al., 1991; Rüthrich et al., 2001). Of course, there are some differences between LTP and kindling-induced potentiation. One of the most striking differences between LTP and kindling is the fact that LTP decays relatively rapidly, within a few 3
hours to a few weeks, whereas kindling is permanent. In addition, LTP typically develops quickly and decays back to baseline relatively rapidly, whereas kindling develops more slowly and is essentially permanent (Cain, et al. 1992; Cain, 1989). Persistence of synaptic potentiation in hippocampus of kindled animals can prevent the induction of new synaptic plasticity and may therefore be involved in learning and memory impairment following kindling (Kemp and Manahan-Vaughan, 2004; Kemp and ManahanVaughan, 2007). Kindling-induced potentiation can remain up to at least 23 (Zhao and Leung, 1991; Zhao and Leung, 1992) or 25 days after the last kindled seizure (Leung and Shen, 1991). Several lines of evidence have demonstrated that low frequency stimulation (LFS; 1-3 Hz) has antiepileptic effects in kindled animal and epileptic humans (Goodman et al., 2005; Mohammad-Zadeh et al., 2007; Yamamoto et al., 2002). It has been shown that LFS can depotentiate the synaptic transmission after LTP induction (Fujii et al., 1991; Larson et al., 1993). Accordingly, it has been suggested that LFS may suppress the kindling-induced synaptic potentiation through mechanisms involved in LTP reversal (Cain, 1989; Mohammad-Zadeh et al., 2007). Our recent findings revealed that application of LFS can improve the kindling-induced impairment in learning and memory in Morris water maze, novel objective recognition task (Esmaeilpour et al., 2017) and Y-maze test (Ghafouri et al., 2016). In addition, the therapeutic effect of LFS in fully kindled animals remains up to one week following its administration (Esmaeilpour et al., 2017). Considering the critical role of LTP in learning and memory (Nabavi et al., 2014), we postulated that improving effect of LFS on seizure-induced impairment in learning and memory is accompanied with its improving effect on LTP induction. Therefore, in
4
the present study we tried to determine if application of LFS in fully kindled animals can lead to a rescue of LTP induction in hippocampal CA1 region and whether this effect is time dependent.
2. Results LTP induction in dorsal hippocampal CA1 area was confirmed by a significant increase in the fEPSP slope (more than 20%) and LTP maintenance was evaluated by recording the fEPSPs for 45 min following primed burst stimulation (PBS). In the slices of control group, application of PBS to the Schaffer collateral in stratum radiatum of dorsal hippocampal CA1 region induced LTP of fEPSP slope that persisted as long as the recording was continued. The experimental protocol used in different experimental groups (with or without LFS application) has been shown in Figure 1. In the first experiment, field potentials were recorded at 48 h following the last kindling stimulation. PBS-induced LTP was impaired in slices of kindled animals. Data analysis showed a significant decrease in LTP induction in the kindled group (named K-48 h group) compared to the control animals (p< 0.001) at 48 h after kindling (Fig. 2). Application of 4 packages of LFS (Fig. 1) in kindled+LFS group (named KLFS- 48 h group) returned the ability of LTP induction to the neuronal circuits of hippocampus so that the mean fEPSP slope significantly increased in the KLFS- 48 h group (p< 0.05) compared to the K-48 h and there was no significant difference between the magnitude of LTP in KLFS-48h and control groups (Fig. 2). It was interesting that although LFS removed the LTP impairment in kindled animals, administration of LFS alone (in non-kindled animals) had inhibitory effect on LTP induction. In this group of animals (LFS48h), application of PBS did not change the fEPSP slope and there was a significant difference between LFS-48h and control groups during post-PBS period (p< 0.001) (Fig. 2). 5
To determine whether the observed effect of LFS is transient or maintains for a long-term following its application, we recorded field potentials at one week after the last kindling stimulation. In kindled animals (K-1 week group) application of PBS was unable to trigger LTP and the increase percentage of fEPSP slope was lower than control group (p<0.001) (Fig. 3). The improving effect of LFS on LTP induction was maintained up to one week after its application so that the LTP magnitude in KLFS-1 week group was significantly higher than the K-1 week group (p<0.05) (Fig. 3). The inhibitory effect of LFS alone on LTP induction was gain observed at one week after its application compared to the control group (p< 0.001( )Fig. 3). The destructive effect of kindling on LTP induction was followed up to 1 month. In this group of animals (K-1 month group) a significant decrease in fEPSP slope was observed after applying PBS compared to control (p<0.001). However, application of 4 packages of LFS after kindling stimulation had no improving effect on LTP induction in KLFS-1 month group and LTP did not induce in this group of animals. In fact, there was no significant difference in the magnitude of LTP between KLFS-1 month and K-1 month groups (Fig. 4). Interestingly, the impairing effect of LFS alone was again observed at 1 month following the last kindling stimulation (Fig. 4). According to above experiments, application of LFS in kindled animals had improving effect on LTP induction up to one week but not one month after kindling stimulations. Therefore, in the next experiment, we tried to determine whether the improving effect of LFS may be detected at two weeks after kindling. Thus, one group of fully kindled animals received 4 packages of LFS and field potential recording were tested 2 weeks after the last kindling stimulation (named KLFS-2 weeks). Obtained results showed that LFS had no improving effect on LTP induction in this group of animals. There was not significant difference in fEPSP slope 6
after PBS application in KLFS-2week group (111.65±2.34 mV/ms) compared to kindle group (103.76±1.56 mV/ms). These results showed that the ameliorating effect of LFS on kindling-induced impairment in LTP maintained only up to one week. Then, we tried to determine whether an increase in the number of LFS packages can extend the duration of its effectiveness. Therefore, in the next experiment in addition to initial 4 packages of LFS (as in the above experiments), another 4 packages of LFS were administered at one week after the last kindling stimulation. The pattern of these LFS packages was exactly similar to the pattern of the initial 4 packages of LFS. In this group of animals, named K2LFS-1 month, field potential recording was done 1 month following the last kindling stimulation. Obtained results showed that the improving effect of LFS on LTP impairment was extended by increasing the number of applied LFS at one-week interval. Application of PBS in this group of animals (n=7) could lead to a significant potentiation in the slope of fEPSP (169.8±6.0 % of baseline; Fig 4). Statistical analysis showed significant difference in fEPSP slope after PBS application between K-1 month and K2LFS-1 month groups (p<0.01). In addition, there was no significant difference between K2LFS-1 month and control groups. To evaluate the effect of kindling and/or LFS on basal synaptic response, input-output (IO) curves were plotted as changes in the slope of fEPSP against the increase in stimulus intensity. The basal synaptic transmission was potentiated at different times (48 h, 1 week and 1 month) after the last kindling stimulation which was detectable as an increase in the fEPSP slope in K-48 h (P<0.001; Fig. 5A), K-1 week (P<0.01; Fig. 5B) and K-1 month (P<0.05; Fig. 5C) compared to control. Administration of LFS clearly suppressed the potentiation effects of
7
kindling on basal synaptic transmission in KLFS-48 h, KLFS-1 week and K2LFS-1 month groups and in these groups the potentiation were similar to control groups (Fig. 5).
Discussion Results of the present study revealed that application of LFS alleviated kindling-induced deficit in LTP induction in CA1 area of the hippocampus. This effect was long-lasting and remained up to one week after the last kindling stimulation and disappeared after two weeks post kindling stimulation. In addition, the effectiveness of LFS can be extended up to one month by increasing the number of LFS applications. Similar to previous reports (Leung and Wu, 2003; Palizvan et al., 2005; Palizvan et al., 2001), kindling lead to a significant impairment of LTP induction in hippocampal CA1 area, so that application of PBS could not produce LTP of fEPSP slope in kindled groups. The impairing effect of kindling on LTP induction was observed even at one month following the last kindling stimulation. In our previous study, a similar time course was observed for kindling-induced impairment in learning and memory in Morris water maze and new objective recognition tests (Esmaeilpour et al., 2017). In fact, in the present study the hippocampal slices were prepared at the same time that the improving effect of LFS on learning and memory was observed in our previous study. On the other hand, the results of the present study, confirmed that LFS application can improve the ability of LTP induction in parallel with improving the learning and memory in fully kindled animals. Several lines of evidence have demonstrated that dorsal hippocampal kindling impairs learning and memory (Da Silva et al., 1986; Kalynchuk and Wintink, 2005; Leung and Shen, 1991; Sutherland et al., 1997). Since the induction of LTP in the hippocampus has an essential 8
role in learning and memory (McNaughton and Morris, 1987; Nabavi et al., 2014; Teyler and Discenna, 1984), therefore, deficits in the hippocampal LTP may be involved in kindlinginduced learning impairments. According to the I-O curves, kindling lead to a significant increase in the slope of fEPSP and this effect was observed even at one month after the last kindling stimulation. Our results are consistent with previous studies showed that the slope of fEPSP increased following kindling (Robinson et al., 1991; Rüthrich et al., 2001; Zhao and Leung, 1991). In fact, kindling can lead to a strong potentiation in synaptic transmission (Cain et al., 1992; Gilbert and Mack, 1990; Racine et al., 1991). It has been shown that , kindling causes long-term enhancement of excitatory synaptic transmission (Douglas and Goddard, 1975) and this kindling-induced plasticity can persist up to 23 days (Zhao and Leung, 1991) or 25 days (Leung and Shen, 1991) after kindling. The long-term persistence of synaptic potentiation may lead to the saturation of all modifiable synapses in a potentiated state, so that synapses could not store further memories (O'Dell and Kandel, 1994). This is the most important reason for the impairment in LTP induction and for deficits in learning and memory after kindling that remain up to 1 month in Morris water maze (Esmaeilpour et al., 2017) and up to 3 weeks (Feasey-Truger et al., 1993) and 4 weeks (Kalynchuk and Fournier, 2009) in radial arm maze performance after kindling. The improving effect of LFS on the kindling-induced impairment in generation of LTP may be through mechanisms involved in depotentiation phenomenon. Depotentiation is the reversal of synaptic strength from potentiation state to pre-LTP levels and LFS (1 Hz) is the most common form of stimuli which results to depotentiation (Chen et al., 2001). According to the I-O curves, application of LFS in the hippocampus of kindled animals reduced the synaptic strength and returned it toward the control situation. Therefore, a mechanism similar to depotentiation 9
may occur to reduce the synaptic potentiation induced by kindling and to increase the ability of information storage in the neural networks (Chen et al., 2001). However, the cellular mechanisms involved in this depotentiation-like phenomenon are unknown. Our previous experiments showed that LFS increases GABAergic currents in fully kindled animals (Asgari et al., 2016). In addition, LFS can reverse both the increment of spontaneous glutamatergic currents and the reduction of spontaneous GABAergic currents in kindled animals (Ghafouri et al., 2016). In our previous study we reported the anticonvulsant action of the LFS pattern which was used in the present study (Esmaeilpour et al., 2017). Considering the possibility of involving a depotentiation-like mechanisms in mediating the observed actions of LFS, and considering the fact that depotentiation is accompanied with a decrease in synaptic transmission, both anticonvulsant and improving effect on the ability of synaptic potentiation may be through the same mechanisms. In a set of previous studies we showed that application of LFS prevented the kindlinginduced potentiation (Jahanshahi et al., 2009; Mohammad-Zadeh et al., 2007; Mohammad-Zadeh et al., 2009). However, it has to be noted that in those studies LFS was applied during kindling stimulation, while in the present study it was administered in fully kindled animals. In the present study, application of LFS alone in non-kindled rats showed significant impairment in LTP compared to control group. This is similar to previous studies which showed that application of LFS alone in non-kindled animals had destructive effect in learning and memory (Esmaeilpour et al., 2017; Yan et al., 2014). This effect may be because of LTD induction following LFS application which can lead to a decrease in synaptic strength and excitatory synaptic transmission in hippocampal CA1 region (Dudek and Bear, 1992).
10
In conclusion, the present data showed that application of LFS in fully kindled animals could return the ability of LTP induction and overcome the kindling-induced impairment in LTP induction in CA1 area of the hippocampus. The durability of this effect can be increased up to one month by increasing the number of applied LFS packages. These changes are completely in parallel to improving effect of LFS on learning and memory in kindled animals.
4. Materials and Methods
4.1 Animals Male Wistar rats (weighing 220–250 g) were used in this study. Animals were caged in groups of four (before surgery) or one (after surgery) with ad libitum access to food and water. They were housed at controlled temperature (23 ± 1 °C) and 12-h light–dark cycle (lights on: 07:00–19:00 h). All experimental protocols and treatments were approved by the ethics committee of Kerman Neuroscience Research Center (ethics code: KNRC-93-53) which is in line with the ‘‘NIH Guide for the Care and Use of Laboratory Animals’’. Efforts were made to minimize both the number of animals used and their suffering.
4.2 Surgery and kindling procedure For stereotaxic surgery, the rats were anesthetized by intra-peritoneal injection of a combination of ketamine (100 mg/kg) and xylazine (12 mg/kg). Animals were implanted with a monopolar recording electrode and bipolar stimulating electrodes which were twisted into a tripolar configuration. This tripolar electrode was positioned in the hippocampal CA1 region (coordinates: A: 2.3 mm; L: 1.7 mm and 2.6 mm below dura) of the right hemisphere. Electrodes 11
(stainless steel, Teflon-coated, 127 μm in diameter, AM-Systems, USA) were insulated except at the cross section of their tips. Another monopolar electrode, which was connected to a small screw, was placed above the left skull surface as ground and differential electrode. All electrodes were connected to pins of a small female plastic socket as a head stage, and fixed to the skull with dental acrylic. Animals had at least 10 days as post-surgical recovery period. After recovery, afterdischarge threshold was determined by applying 3 s monophasic square waves stimulus (1 ms pulse duration) at 50 Hz . Stimulations were initially delivered at 10 µA and then at 5 min intervals, the stimulus intensity was increased in increments of 10 μA until at least 5 s of afterdischarges (ADs) were recorded as previously described (Jahanshahi et al., 2009; Sadegh et al., 2007; Shahpari et al., 2012). The animals were then stimulated daily at the AD threshold intensity in a semi-rapid kindling procedure (12 stimulations/day at 10 min intervals) until three consecutive stage 5 seizures (fully kindled state) were elicited according to Racine scales (Racine, 1972). 4.3 Electrophysiology Hippocampal slices were prepared from rats in all groups. Animals were anesthetized with diethyl ether and decapitated. The brain was quickly dissected and the right hippocampus was separated. Transverse slices (430 µm) were cut with a vibratome (Campden Instruments, UK) in ice-cold artificial cerebrospinal fluid (ACSF). Slices were incubated in a room temperature chamber for at least 60 min. The chamber contained carboxygenated (95% O2, 5% CO2) ACSF, consisting of (in mM): NaCl, 124; NaHCO3, 25; D-glucose, 10; KCl, 4.4; MgCl2 , 2; KH2PO4,H2O, 1.25 and CaCl2, 6H20, 2. Slices were then conveyed on nylon mesh interface
12
chamber, at 32 ◦C, with perfusion ACSF flowing at the rate of 1.5 ml/min. The slices were maintained in this manner for one hour prior to recording. Extracellular field excitatory postsynaptic potentials (fEPSPs) were recorded in the stratum radiatum of the CA1 area of the slice. fEPSPs were evoked by stimulation through a twisted pair of Teflon-coated stainless steel wires placed on the Schaffer collateral fibers. Synaptic responses were recorded using the ACSF-filled micropipettes (4–8 MΩ resistance) placed in the stratum radiatum of hippocampal CA1 area (Fig. 1A). Schaffer collaterals were stimulated at 0.1 Hz by monophasic square pulses (0.1 ms pulse duration) delivered through the bipolar electrode. The Schaffer collateral pathway was stimulated and responses were recorded from the CA1 area to obtain the maximum of fEPSPs slopes. The fEPSP slope was determined as dV/dt during 10% to 90% of the maximum amplitude of fEPSP. The stimulus intensities (input) were gradually increased and fEPSPs (output) were recorded to plot the input-output (I-O) curves. The intensity of the stimulation for baseline recordings was determined at the intensity which evoked a response magnitude of 50% of a maximum slope, i.e. test pulse. The baseline recordings were then obtained by giving a test stimulus every 10 s for 20 min. To achieve LTP, theta pattern primed-burst stimulation (PBS) was used. Each PBS consisted of a single priming pulse, followed 170 ms later by a burst of 10 pulses delivered at 100 Hz. This type of stimulation mimics hippocampal cellular firing patterns accompanied by theta waves. The maintenance phase of the LTP was then recorded for 45 min by applying a test stimulus every 10 s. The mean fEPSPs slope of every 12 sequential traces represents each time point in the graph. Stimulations and recordings were conducted using e-wave and e-pulse instruments and eprobe software version 5.26 (ScienceBeam Co., Tehran, Iran). 13
4.4 Experimental design Five groups of animals were used in the present study including control, sham, control+LFS, kindled, and kindled+LFS. In kindled+LFS group, LFS was applied four times. First and second LFSs were applied immediately and 6 h following the last kindling stimulation respectively. Third and fourth LFSs were applied the next day in a similar manner (i.e. at 6 h interval; Fig. 1). Each LFS consisted of 4 packages at 5 min intervals; each package contained 200 monophasic square wave pulses of 0.1 ms duration at 1 Hz. LFS pattern was achieved according to our preliminary experiments on hippocampal CA1 area (Esmaeilpour et al., 2017; Ghafouri et al., 2016). The intensity of delivered LFS was equal to AD threshold of each kindled rat. Kindled, kindled+LFS and LFS groups were divided into four subgroups, according to the time of hippocampal slice preparation and electrophysiological recording. In these four subgroups hippocampal slices were prepared at 48 h, 1 week, 2 weeks, and 1 month following the last kindling stimulation respectively. In LFS group animals were treated similar to kindled+LFS group, but received only LFS (without kindling stimulations). These animals were also divided into four subgroups and in each subgroup the field potential recordings were done at four times which were exactly similar to the times between LFS and behavioral test in kindled+LFS groups. Another group of animals underwent surgery but did not receive any kind of stimulations and were considered as the sham group. In this group, the elapsed time between surgery and recording was similar to kindled and/or kindled+LFS group. In control group, electrophysiological recordings were done in animals which did not undergo surgery or any kind
14
of stimulations. Seven hippocampal slices from at least 3 rats were used in each experimental group.
4.5 Statistical analysis Data were averaged and expressed as mean ± standard error of the mean (S.E.M.) and accompanied by the number of observations. For each time point of field potential recording during the experiment, average and S.E.M. were calculated from the data on twelve successive evoked responses. A mean value of responses before PBS application was defined as the baseline (100%). Subsequent data were expressed as the percent change from the baseline. The repeated measures two-way ANOVA was used to compare the field potential responses during baseline and LTP induction among the groups. The magnitude of LTP was compared in different experimental groups by one-way ANOVA. Statistically significant differences were evaluated further by Tukey’s post-hoc tests. The probability level interpreted as statistically significant was P<0.05.
Acknowledgement This work was supported financially by Kerman Neuroscience Research Center (Grant No: KNRC/EC/93-53), Kerman, Iran.
References Asgari, A., Semnaian, S., Atapour, N., Shojaei, A., Moradi-Chameh, H., Ghafouri, S., Sheibani, V., Mirnajafi-Zadeh, J., 2016. Low-frequency electrical stimulation enhances the effectiveness of phenobarbital on GABAergic currents in hippocampal slices of kindled rats. Neuroscience. 15
Baudry, M., 1985. Long-term potentiation and kindling: similar biochemical mechanisms? Advances in Neurology. 44, 401-410. Bliss, T.V., Lømo, T., 1973. Long‐lasting potentiation of synaptic transmission in the dentate area of the anaesthetized rabbit following stimulation of the perforant path. The Journal of Physiology. 232, 331-356. Bliss, T.V., Collingridge, G.L., 1993. A synaptic model of memory: long-term potentiation in the hippocampus. Nature. 361, 31-39. Cain, D.P., 1989. Long-term potentiation and kindling: how similar are the mechanisms? Trends in Neurosciences. 12, 6-10. Cain, D.P., Boon, F., Hargreaves, E.L., 1992. Evidence for different neurochemical contributions to long-term potentiation and to kindling and kindling-induced potentiation: role of NMDA and urethane-sensitive mechanisms. Experimental Neurology. 116, 330-338. Cammisuli, S., Murphy, M.P., Ikeda-Douglas, C.J., Balkissoon, V., Holsinger, R.D., Head, E., Michael, M., Racine, R.J., Milgram, N.W., 1997. Effects of extended electrical kindling on exploratory behavior and spatial learning. Behavioural Brain Research. 89, 179-190. Carreño, M., Donaire, A., Rocío Sánchez-Carpintero, M., 2008. Cognitive disorders associated with epilepsy: diagnosis and treatment. The Neurologist. 14, S26-S34. Chen, Y.-L., Huang, C.-C., Hsu, K.-S., 2001. Time-dependent reversal of long-term potentiation by low-frequency stimulation at the hippocampal mossy fiber–CA3 synapses. The Journal of Neuroscience. 21, 3705-3714. Da Silva, F.L., Gorter, J., Wadman, W., 1986. Kindling of the hippocampus induces spatial memory deficits in the rat. Neuroscience Letters. 63, 115-120. Douglas, R.M., Goddard, G.V., 1975. Long-term potentiation of the perforant path-granule cell synapse in the rat hippocampus. Brain Research. 86, 205-215. Dudek, S.M., Bear, M.F., 1992. Homosynaptic long-term depression in area CA1 of hippocampus and effects of N-methyl-D-aspartate receptor blockade. Proceedings of the National Academy of Sciences. 89, 4363-4367. Esmaeilpour, K., Sheibani, V., Shabani, M., Mirnajafi-Zadeh, J., 2017. Effect of low frequency electrical stimulation on seizure-induced short-and long-term impairments in learning and memory in rats. Physiology & Behavior. 168, 112-121. 16
Feasey-Truger, K., Kargl, L., Ten Bruggencate, G., 1993. Differential effects of dentate kindling on working and reference spatial memory in the rat. Neuroscience Letters. 151, 25-28. Fujii, S., Saito, K., Miyakawa, H., Ito, K.-i., Kato, H., 1991. Reversal of long-term potentiation (depotentiation) induced by tetanus stimulation of the input to CA1 neurons of guinea pig hippocampal slices. Brain Research. 555, 112-122. Ghafouri, S., Fathollahi, Y., Javan, M., Shojaei, A., Asgari, A., Mirnajafi-Zadeh, J., 2016. Effect of low frequency stimulation on impaired spontaneous alternation behavior of kindled rats in Y-maze test. Epilepsy research. 126, 37-44. Gilbert, M., Mack, C., 1990. The NMDA antagonist, MK-801, suppresses long-term potentiation, kindling, and kindling-induced potentiation in the perforant path of the unanesthetized rat. Brain research. 519, 89-96. Gilbert, T.H., McNamara, R.K., Corcoran, M.E., 1996. Kindling of hippocampal field CA1 impairs spatial learning and retention in the Morris water maze. Behavioural brain research. 82, 57-66. Goddard, G.V., McIntyre, D.C., Leech, C.K., 1969. A permanent change in brain function resulting from daily electrical stimulation. Experimental neurology. 25, 295-330. Goodman, J.H., Berger, R.E., Tcheng, T.K., 2005. Preemptive Low‐frequency Stimulation Decreases the Incidence of Amygdala‐kindled Seizures. Epilepsia. 46, 1-7. Hannesson, D., Mohapel, P., Corcoran, M., 2001. Dorsal hippocampal kindling selectively impairs spatial learning/short‐term memory. Hippocampus. 11, 275-286. Holmes, G.L., 1991. The long-term effects of seizures on the developing brain: clinical and laboratory issues. Brain and Development. 13, 393-409. Jahanshahi, A., Mirnajafi‐Zadeh, J., Javan, M., Mohammad‐Zadeh, M., Rohani, R., 2009. The antiepileptogenic effect of electrical stimulation at different low frequencies is accompanied with change in adenosine receptors gene expression in rats. Epilepsia. 50, 1768-1779. Kalynchuk, L., Fournier, N., 2009. Interictal anxiety in temporal lobe epilepsy. Encyclopedia of Basic Epilepsy Research. Vol., ed.^eds. San Diego: Elsevier Press/Academic Press. Kalynchuk, L.E., Wintink, A.J., 2005. A potential role for the hippocampus in the expression of kindling-induced fear. In Kindling 6. Vol., ed.^eds. Springer, pp. 285-294. 17
Kemp, A., Manahan-Vaughan, D., 2004. Hippocampal long-term depression and long-term potentiation encode different aspects of novelty acquisition. Proceedings of the National Academy of Sciences of the United States of America. 101, 8192-8197. Kemp, A., Manahan-Vaughan, D., 2007. Hippocampal long-term depression: master or minion in declarative memory processes? Trends in neurosciences. 30, 111-118. Krug, M., Koch, M., Grecksch, G., Schulzeck, K., 1997. Pentylenetetrazol kindling changes the ability to induce potentiation phenomena in the hippocampal CA1 region. Physiology & behavior. 62, 721-727. Larson, J., Xiao, P., Lynch, G., 1993. Reversal of LTP by theta frequency stimulation. Brain research. 600, 97-102. Leung, L.S., Boon, K.A., Kaibara, T., Innis, N.K., 1990. Radial maze performance following hippocampal kindling. Behavioural brain research. 40, 119-129. Leung, L.S., Shen, B., 1991. Hippocampal CA1 evoked response and radial 8-arm maze performance after hippocampal kindling. Brain research. 555, 353-357. Leung, L.S., Wu, C., 2003. Kindling suppresses primed-burst-induced long-term potentiation in hippocampal CA1. Neuroreport. 14, 211-214. Maru, E., Tatsuno, J., Okamoto, J., Ashida, H., 1982. Development and reduction of synaptic potentiation induced by perforant path kindling. Experimental neurology. 78, 409-424. McEachern, J.C., Shaw, C.A., 1996. An alternative to the LTP orthodoxy: a plasticity-pathology continuum model. Brain research reviews. 22, 51-92. McNaughton, B.L., Morris, R.G., 1987. Hippocampal synaptic enhancement and information storage within a distributed memory system. Trends in neurosciences. 10, 408-415. Mohammad-Zadeh, M., Mirnajafi-Zadeh, J., Fathollahi, Y., Javan, M., Ghorbani, P., Sadegh, M., Noorbakhsh, S.M., 2007. Effect of low frequency stimulation of perforant path on kindling rate and synaptic transmission in the dentate gyrus during kindling acquisition in rats. Epilepsy research. 75, 154-161. Mohammad-Zadeh, M., Mirnajafi-Zadeh, J., Fathollahi, Y., Javan, M., Jahanshahi, A., Noorbakhsh, S., Motamedi, F., 2009. The role of adenosine A 1 receptors in mediating the inhibitory effects of low frequency stimulation of perforant path on kindling acquisition in rats. Neuroscience. 158, 1632-1643. 18
Nabavi, S., Fox, R., Proulx, C.D., Lin, J.Y., Tsien, R.Y., Malinow, R., 2014. Engineering a memory with LTD and LTP. Nature. O'Dell, T.J., Kandel, E.R., 1994. Low-frequency stimulation erases LTP through an NMDA receptor-mediated activation of protein phosphatases. Learning & Memory. 1, 129-139. Palizvan, M., Fathollahi, Y., Semnanian, S., 2005. Epileptogenic insult causes a shift in the form of long-term potentiation expression. Neuroscience. 134, 415-423. Palizvan, M.R., Fathollahi, Y., Semnanian, S., Hajezadeh, S., Mirnajafizadh, J., 2001. Differential effects of pentylenetetrazol-kindling on long-term potentiation of population excitatory postsynaptic potentials and population spikes in the CA1 region of rat hippocampus. Brain research. 898, 82-90. Pastalkova, E., Serrano, P., Pinkhasova, D., Wallace, E., Fenton, A.A., Sacktor, T.C., 2006. Storage of spatial information by the maintenance mechanism of LTP. Science. 313, 1141-1144. Racine, R., Newberry, F., Burnham, W., 1975. Post-activation potentiation and the kindling phenomenon. Electroencephalography and clinical neurophysiology. 39, 261-271. Racine, R., 1978. Kindling: the first decade. Neurosurgery. 3, 234-252. Racine, R., Moore, K.-A., Evans, C., 1991. Kindling-induced potentiation in the piriform cortex. Brain research. 556, 218-225. Racine, R.J., 1972. Modification of seizure activity by electrical stimulation: II. Motor seizure. Electroencephalography and clinical neurophysiology. 32, 281-294. Robinson, G.B., Sclabassi, R.J., Berger, T.W., 1991. Kindling-induced potentiation of excitatory and inhibitory inputs to hippocampal dentate granule cells. I. Effects on linear and nonlinear response characteristics. Brain research. 562, 17-25. Rüthrich, H., Grecksch, G., Krug, M., 2001. Development of long-lasting potentiation effects in the dentate gyrus during pentylenetetrazol kindling. International Journal of Developmental Neuroscience. 19, 247-254. Sadegh, M., Mirnajafi-Zadeh, J., Javan, M., Fathollahi, Y., Mohammad-Zadeh, M., Jahanshahi, A., Noorbakhsh, S., 2007. The role of galanin receptors in anticonvulsant effects of lowfrequency stimulation in perforant path–kindled rats. Neuroscience. 150, 396-403.
19
Schiller, Y., Bankirer, Y., 2007. Cellular mechanisms underlying antiepileptic effects of low-and high-frequency electrical stimulation in acute epilepsy in neocortical brain slices in vitro. Journal of neurophysiology. 97, 1887-1902. Shahpari, M., Mirnajafi-Zadeh, J., Firoozabadi, S.M.P., Yadollahpour, A., 2012. Effect of lowfrequency electrical stimulation parameters on its anticonvulsant action during rapid perforant path kindling in rat. Epilepsy research. 99, 69-77. Soderling, T.R., Derkach, V.A., 2000. Postsynaptic protein phosphorylation and LTP. Trends in neurosciences. 23, 75-80. Sutherland, R.J., Leung, L., Weisend, M.P., Schlife, J., McDonald, R.J., 1997. An evaluation of the effect of partial hippocampal kindling on place navigation by rats in the Morris water task. Psychobiology. 25, 126-132. Sutula, T., Steward, O., 1986. Quantitative analysis of synaptic potentiation during kindling of the perforant path. Journal of neurophysiology. 56, 732-746. Teyler, T.J., Discenna, P., 1984. Long-term potentiation as a candidate mnemonic device. Brain research reviews. 7, 15-28. Teyler, T.J., DiScenna, P., 1985. The role of hippocampus in memory: a hypothesis. Neuroscience & Biobehavioral Reviews. 9, 377-389. Volcy, G.M., 2003. [Mesial temporal lobe epilepsy: its physiopathology, clinical characteristics, treatment and prognosis]. Revista de neurologia. 38, 663-667. Yamamoto, J., Ikeda, A., Satow, T., Takeshita, K., Takayama, M., Matsuhashi, M., Matsumoto, R., Ohara, S., Mikuni, N., Takahashi, J., 2002. Low‐frequency Electric Cortical Stimulation Has an Inhibitory Effect on Epileptic Focus in Mesial Temporal Lobe Epilepsy. Epilepsia. 43, 491-495. Yan, W.-W., Wang, C.-Y., Zeng, J., Liu, Q.-Y., Xu, S.-T., Liu, W.-X., Xiao, P., Li, C.-H., 2014. Low-frequency stimulation of dorsal norephinephrine bundle reverses behavioral longterm potentiation and learning performance in rats. Neuroscience. 265, 238-244. Zhao, D., Leung, L.S., 1991. Effects of hippocampal kindling on paired-pulse response in CA1 in vitro. Brain research. 564, 220-229. Zhao, D., Leung, L.S., 1992. Hippocampal kindling induced paired-pulse depression in the dentate gyrus and paired-pulse facilitation in CA3. Brain research. 582, 163-167. 20
21
Figures legends Fig. 1. Time-line diagram showing the protocol used for LFS application in animals of kindled+LFS group.
Fig. 2. Effect low-frequency stimulation (LFS) on kindling-induced impairment in LTP induction in CA1 area of the hippocampus at 48h after kindling stimulation. (A) schematic figure showing the place of stimulating and recording electrodes in hippocampal slice (DG: dentate gyrus; sch: Schaffer collaterals). (B) Sample records of field excitatory post-synaptic potentials (fEPSP) recording at 48 h after the last kindling stimulation in control, kindled (K-48 h), kindled+LFS (KLFS-48 h) and LFS (LFS-48 h) groups. Numbers 1 and 2 show the time of fEPSP recording according to the time-course graphs in C. (C) Time-course of fEPSP before and after application of primed-burst stimulation (PBS) in experimental groups. Data are plotted as the average percentage change from baseline responses. Each point shows the mean ± SEM. Application of PBS could not induce LTP in K-48 h group. However, application of LFS (in KLFS-48 h group) returned the ability of LTP induction. Application of LFS alone had also inhibitory effect on LTP induction. (D) Bar diagrams showing the changes in percentage of fEPSP slope after application of PBS (from 0 to 45 min) in different experimental groups. Each bar is mean ± SEM . *** p<0.001 compared to control group. # p<0.05 compared to K-48 h group (n=7 in all groups).
Fig. 3. Effect low-frequency stimulation (LFS) on kindling-induced impairment in LTP induction in CA1 area of the hippocampus at one week after kindling stimulation. (A) Sample records of field excitatory post-synaptic potentials (fEPSP) recording at one week after the last 22
kindling stimulation in control, kindled (K-1 week), kindled+LFS (KLFS-1 week) and LFS (LFS-1 week) groups. Numbers 1 and 2 show the time of fEPSP recording according to the timecourse graphs in B. (B) Time-course of fEPSP before and after application of primed-burst stimulation (PBS) in experimental groups. Data are plotted as the average percentage change from baseline responses. Each point shows the mean ± SEM. Application of PBS did not induce LTP in K-1 week group. However, application of LFS (in KLFS-1 week group) returned the ability of LTP induction. Application of LFS alone had also inhibitory effect on LTP induction. (C) Bar diagrams showing the changes in percentage of fEPSP slope after application of PBS (from 0 to 45 min) in different experimental groups. Each bar is mean ± SEM. *** p<0.001 compared to control group.
#
p<0.05 compared to K-1 week group (n=7 in all groups).
Fig. 4. Effect low-frequency stimulation (LFS) on kindling-induced impairment in LTP induction in CA1 area of the hippocampus at one month after kindling stimulation. (A) Sample records of field excitatory post-synaptic potentials (fEPSP) recording at one month after the last kindling stimulation in control, kindled (K-1 week), kindled+LFS (KLFS-1 week), kindled+2 LFS (K2LFS-1 month) and LFS (LFS-1 month) groups. Numbers 1 and 2 show the time of fEPSP recording according to the time-course graphs in B. (B) Time-course of fEPSP before and after application of primed-burst stimulation (PBS) in experimental groups. Data are plotted as the average percentage change from baseline responses. Each point shows the mean ± SEM. Application of PBS could not induce LTP in K-1 month and KLFS-1 month groups. However, application of additional LFS packages at one week after the first packages (see text) (K2LFS-1 month group) returned the ability of LTP induction in hippocampal slices prepared one month after the last kindling stimulation. Application of LFS alone had also inhibitory effect on LTP 23
induction. (C) Bar diagrams showing the changes in percentage of fEPSP slope after application of PBS (from 0 to 45 min) in different experimental groups. Each bar is mean ± SEM. *** p<0.001 compared to control group. ## p<0.01 compared to K-1 month group (n=7 in all groups).
Fig. 5. Input-output (I-O) curves were acquired according to the slope of field excitatory postsynaptic potential (fEPSP) in response to different stimulus intensities in the CA1 area of the hippocampus. I-O curves were drawn at 48 h (A), 1 week (B) and 1 month (C) after the last kindling stimulation in different experimental groups. There was a significant left-side shift in IO curve of kindled animals at 48 h (K-48 h group), 1 week (K-1 week) and 1 month (K-1 month) following the last kindling stimulation. Application of LFS in kindled animals completely returned this left-side shift toward control curve in hippocampal slices prepared at 48 h (KLFS48 h) and one week (KLFS- 1 week), but not one month (KLFS-1 month) after the last kindling stimulation. However, application of additional LFS packages at one week after the first packages (see text) could completely return the I-O curve toward control situation at one-month after kindling (K2LFS-1 month group).
24
Highlights: Long-term potentiation (LTP) cannot be induced in hippocampus up to 1 month after kindling. Low-frequency stimulation (LFS) alleviated kindling-induced deficit in LTP induction. Improving effect of LFS on LTP induction remained up to one week in kindled animals. LFS effects extended up to one month by increasing the number of its applications.
25
26
27
28
29
30
S0006-8993(17)30196-8 http://dx.doi.org/10.1016/j.brainres.2017.05.007 BRES 45356
To appear in:
Brain Research
Received Date: Revised Date: Accepted Date:
24 December 2016 28 April 2017 8 May 2017
Please cite this article as: K. Esmaeilpour, V. Sheibani, M. Shabani, J. Mirnajafi-Zadeh, Low frequency electrical stimulation has time dependent improving effect on kindling-induced impairment in long-term potentiation in rats, Brain Research (2017), doi: http://dx.doi.org/10.1016/j.brainres.2017.05.007
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Low frequency electrical stimulation has time dependent improving effect on kindling-induced impairment in long-term potentiation in rats
Khadijeh Esmaeilpour1, Vahid Sheibani1, Mohammad Shabani1, Javad Mirnajafi-Zadeh1,2*
1- Neuroscience Research Center, Institute of Neuropharmacology, Kerman University of Medical Sciences, Kerman, Iran 2- Department of Physiology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
*Corresponding author: Javad Mirnajafi-Zadeh, Department of Physiology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, PO Box: 14115-331, Iran, Tel: +98-21 82883865, Fax: +98-21 82884825, E-mail: [email protected]
1
Abstract Application of low-frequency stimulation (LFS) can improve learning and memory in kindled animals (Ghafouri et al., 2016). Considering the important role of long-term potentiation (LTP) in learning and memory, in the present study the effectiveness of LFS on kindling-induced impairment in LTP induction was investigated in hippocampal CA1 area at different times post kindling stimulations. Animals were kindled via electrical stimulation of hippocampal CA1 area in a semi-rapid manner (12 stimulations per day). One group of animals received four trials of LFS at 30 s, 6 h, 24 h, and 30 h following the last kindling stimulation. Each LFS consisted of 4 packages at 5 min intervals; each package contained 200 monophasic square wave pulses of 0.1 ms duration at 1 Hz. The kindled, kindled+LFS and LFS groups were divided into four subgroups in which hippocampal slices were prepared at 48 h, 1 week, 2 weeks, and 1 month following the last kindling stimulation respectively. Extracellular evoked field excitatory postsynaptic potentials (fEPSPs) were recorded in the stratum radiatum of the CA1 area of the slice. Obtained results showed that LTP was not induced in kindled animals. However, application of LFS overcame the kindling-induced impairment in LTP generation in CA1 area of the hippocampus. This improving effect remained up to one week after the last kindling stimulation and extended to one month by increasing the number of applied LFS packages.
Keywords: Seizure; Low frequency stimulation; Long-term potentiation; Hippocampus
2
1. Introduction Temporal lobe epilepsy is one of the most prevalent neurological disorders (Schiller and Bankirer, 2007; Volcy, 2003). Impairment in learning and memory has been reported in patients with epilepsy (Carreño et al., 2008; Holmes, 1991). Kindling is an experimental animal model for temporal lobe epilepsy which can be induced by daily low-intensity electrical stimulation of limbic areas such as the hippocampus (Goddard et al., 1969; Racine, 1978). Similar to epileptic patients, kindled animals show impairment in spatial learning and memory which maybe a result of damage to neuronal function in the hippocampus (Cammisuli et al., 1997; Da Silva et al., 1986; Gilbert et al., 1996; Hannesson et al., 2001; Leung et al., 1990). Long-term potentiation (LTP) in the hippocampus is a form of synaptic plasticity which has been considered as the neuronal mechanism involved in learning and memory (Bliss and Lømo, 1973; Bliss and Collingridge, 1993; Nabavi et al., 2014; Pastalkova et al., 2006; Soderling and Derkach, 2000; Teyler and DiScenna, 1985). Kindling phenomenon is accompanied with synaptic potentiation and there are many similarities between kindling-induced potentiation and LTP (Krug et al., 1997; McEachern and Shaw, 1996). Accordingly, a possible mechanism of kindling epileptogenesis is long-lasting enhancement of excitatory synapses (Baudry, 1985; Douglas and Goddard, 1975; Maru et al., 1982; Racine et al., 1975; Sutula and Steward, 1986). Many studies have indicated that kindled seizures can lead to synaptic potentiation which is accompanied with an increase in slope of field excitatory postsynaptic potential (fEPSP) and amplitude of population spike (PS) (Jahanshahi et al., 2009; Mohammad-Zadeh et al., 2007; Mohammad-Zadeh et al., 2009; Robinson et al., 1991; Rüthrich et al., 2001). Of course, there are some differences between LTP and kindling-induced potentiation. One of the most striking differences between LTP and kindling is the fact that LTP decays relatively rapidly, within a few 3
hours to a few weeks, whereas kindling is permanent. In addition, LTP typically develops quickly and decays back to baseline relatively rapidly, whereas kindling develops more slowly and is essentially permanent (Cain, et al. 1992; Cain, 1989). Persistence of synaptic potentiation in hippocampus of kindled animals can prevent the induction of new synaptic plasticity and may therefore be involved in learning and memory impairment following kindling (Kemp and Manahan-Vaughan, 2004; Kemp and ManahanVaughan, 2007). Kindling-induced potentiation can remain up to at least 23 (Zhao and Leung, 1991; Zhao and Leung, 1992) or 25 days after the last kindled seizure (Leung and Shen, 1991). Several lines of evidence have demonstrated that low frequency stimulation (LFS; 1-3 Hz) has antiepileptic effects in kindled animal and epileptic humans (Goodman et al., 2005; Mohammad-Zadeh et al., 2007; Yamamoto et al., 2002). It has been shown that LFS can depotentiate the synaptic transmission after LTP induction (Fujii et al., 1991; Larson et al., 1993). Accordingly, it has been suggested that LFS may suppress the kindling-induced synaptic potentiation through mechanisms involved in LTP reversal (Cain, 1989; Mohammad-Zadeh et al., 2007). Our recent findings revealed that application of LFS can improve the kindling-induced impairment in learning and memory in Morris water maze, novel objective recognition task (Esmaeilpour et al., 2017) and Y-maze test (Ghafouri et al., 2016). In addition, the therapeutic effect of LFS in fully kindled animals remains up to one week following its administration (Esmaeilpour et al., 2017). Considering the critical role of LTP in learning and memory (Nabavi et al., 2014), we postulated that improving effect of LFS on seizure-induced impairment in learning and memory is accompanied with its improving effect on LTP induction. Therefore, in
4
the present study we tried to determine if application of LFS in fully kindled animals can lead to a rescue of LTP induction in hippocampal CA1 region and whether this effect is time dependent.
2. Results LTP induction in dorsal hippocampal CA1 area was confirmed by a significant increase in the fEPSP slope (more than 20%) and LTP maintenance was evaluated by recording the fEPSPs for 45 min following primed burst stimulation (PBS). In the slices of control group, application of PBS to the Schaffer collateral in stratum radiatum of dorsal hippocampal CA1 region induced LTP of fEPSP slope that persisted as long as the recording was continued. The experimental protocol used in different experimental groups (with or without LFS application) has been shown in Figure 1. In the first experiment, field potentials were recorded at 48 h following the last kindling stimulation. PBS-induced LTP was impaired in slices of kindled animals. Data analysis showed a significant decrease in LTP induction in the kindled group (named K-48 h group) compared to the control animals (p< 0.001) at 48 h after kindling (Fig. 2). Application of 4 packages of LFS (Fig. 1) in kindled+LFS group (named KLFS- 48 h group) returned the ability of LTP induction to the neuronal circuits of hippocampus so that the mean fEPSP slope significantly increased in the KLFS- 48 h group (p< 0.05) compared to the K-48 h and there was no significant difference between the magnitude of LTP in KLFS-48h and control groups (Fig. 2). It was interesting that although LFS removed the LTP impairment in kindled animals, administration of LFS alone (in non-kindled animals) had inhibitory effect on LTP induction. In this group of animals (LFS48h), application of PBS did not change the fEPSP slope and there was a significant difference between LFS-48h and control groups during post-PBS period (p< 0.001) (Fig. 2). 5
To determine whether the observed effect of LFS is transient or maintains for a long-term following its application, we recorded field potentials at one week after the last kindling stimulation. In kindled animals (K-1 week group) application of PBS was unable to trigger LTP and the increase percentage of fEPSP slope was lower than control group (p<0.001) (Fig. 3). The improving effect of LFS on LTP induction was maintained up to one week after its application so that the LTP magnitude in KLFS-1 week group was significantly higher than the K-1 week group (p<0.05) (Fig. 3). The inhibitory effect of LFS alone on LTP induction was gain observed at one week after its application compared to the control group (p< 0.001( )Fig. 3). The destructive effect of kindling on LTP induction was followed up to 1 month. In this group of animals (K-1 month group) a significant decrease in fEPSP slope was observed after applying PBS compared to control (p<0.001). However, application of 4 packages of LFS after kindling stimulation had no improving effect on LTP induction in KLFS-1 month group and LTP did not induce in this group of animals. In fact, there was no significant difference in the magnitude of LTP between KLFS-1 month and K-1 month groups (Fig. 4). Interestingly, the impairing effect of LFS alone was again observed at 1 month following the last kindling stimulation (Fig. 4). According to above experiments, application of LFS in kindled animals had improving effect on LTP induction up to one week but not one month after kindling stimulations. Therefore, in the next experiment, we tried to determine whether the improving effect of LFS may be detected at two weeks after kindling. Thus, one group of fully kindled animals received 4 packages of LFS and field potential recording were tested 2 weeks after the last kindling stimulation (named KLFS-2 weeks). Obtained results showed that LFS had no improving effect on LTP induction in this group of animals. There was not significant difference in fEPSP slope 6
after PBS application in KLFS-2week group (111.65±2.34 mV/ms) compared to kindle group (103.76±1.56 mV/ms). These results showed that the ameliorating effect of LFS on kindling-induced impairment in LTP maintained only up to one week. Then, we tried to determine whether an increase in the number of LFS packages can extend the duration of its effectiveness. Therefore, in the next experiment in addition to initial 4 packages of LFS (as in the above experiments), another 4 packages of LFS were administered at one week after the last kindling stimulation. The pattern of these LFS packages was exactly similar to the pattern of the initial 4 packages of LFS. In this group of animals, named K2LFS-1 month, field potential recording was done 1 month following the last kindling stimulation. Obtained results showed that the improving effect of LFS on LTP impairment was extended by increasing the number of applied LFS at one-week interval. Application of PBS in this group of animals (n=7) could lead to a significant potentiation in the slope of fEPSP (169.8±6.0 % of baseline; Fig 4). Statistical analysis showed significant difference in fEPSP slope after PBS application between K-1 month and K2LFS-1 month groups (p<0.01). In addition, there was no significant difference between K2LFS-1 month and control groups. To evaluate the effect of kindling and/or LFS on basal synaptic response, input-output (IO) curves were plotted as changes in the slope of fEPSP against the increase in stimulus intensity. The basal synaptic transmission was potentiated at different times (48 h, 1 week and 1 month) after the last kindling stimulation which was detectable as an increase in the fEPSP slope in K-48 h (P<0.001; Fig. 5A), K-1 week (P<0.01; Fig. 5B) and K-1 month (P<0.05; Fig. 5C) compared to control. Administration of LFS clearly suppressed the potentiation effects of
7
kindling on basal synaptic transmission in KLFS-48 h, KLFS-1 week and K2LFS-1 month groups and in these groups the potentiation were similar to control groups (Fig. 5).
Discussion Results of the present study revealed that application of LFS alleviated kindling-induced deficit in LTP induction in CA1 area of the hippocampus. This effect was long-lasting and remained up to one week after the last kindling stimulation and disappeared after two weeks post kindling stimulation. In addition, the effectiveness of LFS can be extended up to one month by increasing the number of LFS applications. Similar to previous reports (Leung and Wu, 2003; Palizvan et al., 2005; Palizvan et al., 2001), kindling lead to a significant impairment of LTP induction in hippocampal CA1 area, so that application of PBS could not produce LTP of fEPSP slope in kindled groups. The impairing effect of kindling on LTP induction was observed even at one month following the last kindling stimulation. In our previous study, a similar time course was observed for kindling-induced impairment in learning and memory in Morris water maze and new objective recognition tests (Esmaeilpour et al., 2017). In fact, in the present study the hippocampal slices were prepared at the same time that the improving effect of LFS on learning and memory was observed in our previous study. On the other hand, the results of the present study, confirmed that LFS application can improve the ability of LTP induction in parallel with improving the learning and memory in fully kindled animals. Several lines of evidence have demonstrated that dorsal hippocampal kindling impairs learning and memory (Da Silva et al., 1986; Kalynchuk and Wintink, 2005; Leung and Shen, 1991; Sutherland et al., 1997). Since the induction of LTP in the hippocampus has an essential 8
role in learning and memory (McNaughton and Morris, 1987; Nabavi et al., 2014; Teyler and Discenna, 1984), therefore, deficits in the hippocampal LTP may be involved in kindlinginduced learning impairments. According to the I-O curves, kindling lead to a significant increase in the slope of fEPSP and this effect was observed even at one month after the last kindling stimulation. Our results are consistent with previous studies showed that the slope of fEPSP increased following kindling (Robinson et al., 1991; Rüthrich et al., 2001; Zhao and Leung, 1991). In fact, kindling can lead to a strong potentiation in synaptic transmission (Cain et al., 1992; Gilbert and Mack, 1990; Racine et al., 1991). It has been shown that , kindling causes long-term enhancement of excitatory synaptic transmission (Douglas and Goddard, 1975) and this kindling-induced plasticity can persist up to 23 days (Zhao and Leung, 1991) or 25 days (Leung and Shen, 1991) after kindling. The long-term persistence of synaptic potentiation may lead to the saturation of all modifiable synapses in a potentiated state, so that synapses could not store further memories (O'Dell and Kandel, 1994). This is the most important reason for the impairment in LTP induction and for deficits in learning and memory after kindling that remain up to 1 month in Morris water maze (Esmaeilpour et al., 2017) and up to 3 weeks (Feasey-Truger et al., 1993) and 4 weeks (Kalynchuk and Fournier, 2009) in radial arm maze performance after kindling. The improving effect of LFS on the kindling-induced impairment in generation of LTP may be through mechanisms involved in depotentiation phenomenon. Depotentiation is the reversal of synaptic strength from potentiation state to pre-LTP levels and LFS (1 Hz) is the most common form of stimuli which results to depotentiation (Chen et al., 2001). According to the I-O curves, application of LFS in the hippocampus of kindled animals reduced the synaptic strength and returned it toward the control situation. Therefore, a mechanism similar to depotentiation 9
may occur to reduce the synaptic potentiation induced by kindling and to increase the ability of information storage in the neural networks (Chen et al., 2001). However, the cellular mechanisms involved in this depotentiation-like phenomenon are unknown. Our previous experiments showed that LFS increases GABAergic currents in fully kindled animals (Asgari et al., 2016). In addition, LFS can reverse both the increment of spontaneous glutamatergic currents and the reduction of spontaneous GABAergic currents in kindled animals (Ghafouri et al., 2016). In our previous study we reported the anticonvulsant action of the LFS pattern which was used in the present study (Esmaeilpour et al., 2017). Considering the possibility of involving a depotentiation-like mechanisms in mediating the observed actions of LFS, and considering the fact that depotentiation is accompanied with a decrease in synaptic transmission, both anticonvulsant and improving effect on the ability of synaptic potentiation may be through the same mechanisms. In a set of previous studies we showed that application of LFS prevented the kindlinginduced potentiation (Jahanshahi et al., 2009; Mohammad-Zadeh et al., 2007; Mohammad-Zadeh et al., 2009). However, it has to be noted that in those studies LFS was applied during kindling stimulation, while in the present study it was administered in fully kindled animals. In the present study, application of LFS alone in non-kindled rats showed significant impairment in LTP compared to control group. This is similar to previous studies which showed that application of LFS alone in non-kindled animals had destructive effect in learning and memory (Esmaeilpour et al., 2017; Yan et al., 2014). This effect may be because of LTD induction following LFS application which can lead to a decrease in synaptic strength and excitatory synaptic transmission in hippocampal CA1 region (Dudek and Bear, 1992).
10
In conclusion, the present data showed that application of LFS in fully kindled animals could return the ability of LTP induction and overcome the kindling-induced impairment in LTP induction in CA1 area of the hippocampus. The durability of this effect can be increased up to one month by increasing the number of applied LFS packages. These changes are completely in parallel to improving effect of LFS on learning and memory in kindled animals.
4. Materials and Methods
4.1 Animals Male Wistar rats (weighing 220–250 g) were used in this study. Animals were caged in groups of four (before surgery) or one (after surgery) with ad libitum access to food and water. They were housed at controlled temperature (23 ± 1 °C) and 12-h light–dark cycle (lights on: 07:00–19:00 h). All experimental protocols and treatments were approved by the ethics committee of Kerman Neuroscience Research Center (ethics code: KNRC-93-53) which is in line with the ‘‘NIH Guide for the Care and Use of Laboratory Animals’’. Efforts were made to minimize both the number of animals used and their suffering.
4.2 Surgery and kindling procedure For stereotaxic surgery, the rats were anesthetized by intra-peritoneal injection of a combination of ketamine (100 mg/kg) and xylazine (12 mg/kg). Animals were implanted with a monopolar recording electrode and bipolar stimulating electrodes which were twisted into a tripolar configuration. This tripolar electrode was positioned in the hippocampal CA1 region (coordinates: A: 2.3 mm; L: 1.7 mm and 2.6 mm below dura) of the right hemisphere. Electrodes 11
(stainless steel, Teflon-coated, 127 μm in diameter, AM-Systems, USA) were insulated except at the cross section of their tips. Another monopolar electrode, which was connected to a small screw, was placed above the left skull surface as ground and differential electrode. All electrodes were connected to pins of a small female plastic socket as a head stage, and fixed to the skull with dental acrylic. Animals had at least 10 days as post-surgical recovery period. After recovery, afterdischarge threshold was determined by applying 3 s monophasic square waves stimulus (1 ms pulse duration) at 50 Hz . Stimulations were initially delivered at 10 µA and then at 5 min intervals, the stimulus intensity was increased in increments of 10 μA until at least 5 s of afterdischarges (ADs) were recorded as previously described (Jahanshahi et al., 2009; Sadegh et al., 2007; Shahpari et al., 2012). The animals were then stimulated daily at the AD threshold intensity in a semi-rapid kindling procedure (12 stimulations/day at 10 min intervals) until three consecutive stage 5 seizures (fully kindled state) were elicited according to Racine scales (Racine, 1972). 4.3 Electrophysiology Hippocampal slices were prepared from rats in all groups. Animals were anesthetized with diethyl ether and decapitated. The brain was quickly dissected and the right hippocampus was separated. Transverse slices (430 µm) were cut with a vibratome (Campden Instruments, UK) in ice-cold artificial cerebrospinal fluid (ACSF). Slices were incubated in a room temperature chamber for at least 60 min. The chamber contained carboxygenated (95% O2, 5% CO2) ACSF, consisting of (in mM): NaCl, 124; NaHCO3, 25; D-glucose, 10; KCl, 4.4; MgCl2 , 2; KH2PO4,H2O, 1.25 and CaCl2, 6H20, 2. Slices were then conveyed on nylon mesh interface
12
chamber, at 32 ◦C, with perfusion ACSF flowing at the rate of 1.5 ml/min. The slices were maintained in this manner for one hour prior to recording. Extracellular field excitatory postsynaptic potentials (fEPSPs) were recorded in the stratum radiatum of the CA1 area of the slice. fEPSPs were evoked by stimulation through a twisted pair of Teflon-coated stainless steel wires placed on the Schaffer collateral fibers. Synaptic responses were recorded using the ACSF-filled micropipettes (4–8 MΩ resistance) placed in the stratum radiatum of hippocampal CA1 area (Fig. 1A). Schaffer collaterals were stimulated at 0.1 Hz by monophasic square pulses (0.1 ms pulse duration) delivered through the bipolar electrode. The Schaffer collateral pathway was stimulated and responses were recorded from the CA1 area to obtain the maximum of fEPSPs slopes. The fEPSP slope was determined as dV/dt during 10% to 90% of the maximum amplitude of fEPSP. The stimulus intensities (input) were gradually increased and fEPSPs (output) were recorded to plot the input-output (I-O) curves. The intensity of the stimulation for baseline recordings was determined at the intensity which evoked a response magnitude of 50% of a maximum slope, i.e. test pulse. The baseline recordings were then obtained by giving a test stimulus every 10 s for 20 min. To achieve LTP, theta pattern primed-burst stimulation (PBS) was used. Each PBS consisted of a single priming pulse, followed 170 ms later by a burst of 10 pulses delivered at 100 Hz. This type of stimulation mimics hippocampal cellular firing patterns accompanied by theta waves. The maintenance phase of the LTP was then recorded for 45 min by applying a test stimulus every 10 s. The mean fEPSPs slope of every 12 sequential traces represents each time point in the graph. Stimulations and recordings were conducted using e-wave and e-pulse instruments and eprobe software version 5.26 (ScienceBeam Co., Tehran, Iran). 13
4.4 Experimental design Five groups of animals were used in the present study including control, sham, control+LFS, kindled, and kindled+LFS. In kindled+LFS group, LFS was applied four times. First and second LFSs were applied immediately and 6 h following the last kindling stimulation respectively. Third and fourth LFSs were applied the next day in a similar manner (i.e. at 6 h interval; Fig. 1). Each LFS consisted of 4 packages at 5 min intervals; each package contained 200 monophasic square wave pulses of 0.1 ms duration at 1 Hz. LFS pattern was achieved according to our preliminary experiments on hippocampal CA1 area (Esmaeilpour et al., 2017; Ghafouri et al., 2016). The intensity of delivered LFS was equal to AD threshold of each kindled rat. Kindled, kindled+LFS and LFS groups were divided into four subgroups, according to the time of hippocampal slice preparation and electrophysiological recording. In these four subgroups hippocampal slices were prepared at 48 h, 1 week, 2 weeks, and 1 month following the last kindling stimulation respectively. In LFS group animals were treated similar to kindled+LFS group, but received only LFS (without kindling stimulations). These animals were also divided into four subgroups and in each subgroup the field potential recordings were done at four times which were exactly similar to the times between LFS and behavioral test in kindled+LFS groups. Another group of animals underwent surgery but did not receive any kind of stimulations and were considered as the sham group. In this group, the elapsed time between surgery and recording was similar to kindled and/or kindled+LFS group. In control group, electrophysiological recordings were done in animals which did not undergo surgery or any kind
14
of stimulations. Seven hippocampal slices from at least 3 rats were used in each experimental group.
4.5 Statistical analysis Data were averaged and expressed as mean ± standard error of the mean (S.E.M.) and accompanied by the number of observations. For each time point of field potential recording during the experiment, average and S.E.M. were calculated from the data on twelve successive evoked responses. A mean value of responses before PBS application was defined as the baseline (100%). Subsequent data were expressed as the percent change from the baseline. The repeated measures two-way ANOVA was used to compare the field potential responses during baseline and LTP induction among the groups. The magnitude of LTP was compared in different experimental groups by one-way ANOVA. Statistically significant differences were evaluated further by Tukey’s post-hoc tests. The probability level interpreted as statistically significant was P<0.05.
Acknowledgement This work was supported financially by Kerman Neuroscience Research Center (Grant No: KNRC/EC/93-53), Kerman, Iran.
References Asgari, A., Semnaian, S., Atapour, N., Shojaei, A., Moradi-Chameh, H., Ghafouri, S., Sheibani, V., Mirnajafi-Zadeh, J., 2016. Low-frequency electrical stimulation enhances the effectiveness of phenobarbital on GABAergic currents in hippocampal slices of kindled rats. Neuroscience. 15
Baudry, M., 1985. Long-term potentiation and kindling: similar biochemical mechanisms? Advances in Neurology. 44, 401-410. Bliss, T.V., Lømo, T., 1973. Long‐lasting potentiation of synaptic transmission in the dentate area of the anaesthetized rabbit following stimulation of the perforant path. The Journal of Physiology. 232, 331-356. Bliss, T.V., Collingridge, G.L., 1993. A synaptic model of memory: long-term potentiation in the hippocampus. Nature. 361, 31-39. Cain, D.P., 1989. Long-term potentiation and kindling: how similar are the mechanisms? Trends in Neurosciences. 12, 6-10. Cain, D.P., Boon, F., Hargreaves, E.L., 1992. Evidence for different neurochemical contributions to long-term potentiation and to kindling and kindling-induced potentiation: role of NMDA and urethane-sensitive mechanisms. Experimental Neurology. 116, 330-338. Cammisuli, S., Murphy, M.P., Ikeda-Douglas, C.J., Balkissoon, V., Holsinger, R.D., Head, E., Michael, M., Racine, R.J., Milgram, N.W., 1997. Effects of extended electrical kindling on exploratory behavior and spatial learning. Behavioural Brain Research. 89, 179-190. Carreño, M., Donaire, A., Rocío Sánchez-Carpintero, M., 2008. Cognitive disorders associated with epilepsy: diagnosis and treatment. The Neurologist. 14, S26-S34. Chen, Y.-L., Huang, C.-C., Hsu, K.-S., 2001. Time-dependent reversal of long-term potentiation by low-frequency stimulation at the hippocampal mossy fiber–CA3 synapses. The Journal of Neuroscience. 21, 3705-3714. Da Silva, F.L., Gorter, J., Wadman, W., 1986. Kindling of the hippocampus induces spatial memory deficits in the rat. Neuroscience Letters. 63, 115-120. Douglas, R.M., Goddard, G.V., 1975. Long-term potentiation of the perforant path-granule cell synapse in the rat hippocampus. Brain Research. 86, 205-215. Dudek, S.M., Bear, M.F., 1992. Homosynaptic long-term depression in area CA1 of hippocampus and effects of N-methyl-D-aspartate receptor blockade. Proceedings of the National Academy of Sciences. 89, 4363-4367. Esmaeilpour, K., Sheibani, V., Shabani, M., Mirnajafi-Zadeh, J., 2017. Effect of low frequency electrical stimulation on seizure-induced short-and long-term impairments in learning and memory in rats. Physiology & Behavior. 168, 112-121. 16
Feasey-Truger, K., Kargl, L., Ten Bruggencate, G., 1993. Differential effects of dentate kindling on working and reference spatial memory in the rat. Neuroscience Letters. 151, 25-28. Fujii, S., Saito, K., Miyakawa, H., Ito, K.-i., Kato, H., 1991. Reversal of long-term potentiation (depotentiation) induced by tetanus stimulation of the input to CA1 neurons of guinea pig hippocampal slices. Brain Research. 555, 112-122. Ghafouri, S., Fathollahi, Y., Javan, M., Shojaei, A., Asgari, A., Mirnajafi-Zadeh, J., 2016. Effect of low frequency stimulation on impaired spontaneous alternation behavior of kindled rats in Y-maze test. Epilepsy research. 126, 37-44. Gilbert, M., Mack, C., 1990. The NMDA antagonist, MK-801, suppresses long-term potentiation, kindling, and kindling-induced potentiation in the perforant path of the unanesthetized rat. Brain research. 519, 89-96. Gilbert, T.H., McNamara, R.K., Corcoran, M.E., 1996. Kindling of hippocampal field CA1 impairs spatial learning and retention in the Morris water maze. Behavioural brain research. 82, 57-66. Goddard, G.V., McIntyre, D.C., Leech, C.K., 1969. A permanent change in brain function resulting from daily electrical stimulation. Experimental neurology. 25, 295-330. Goodman, J.H., Berger, R.E., Tcheng, T.K., 2005. Preemptive Low‐frequency Stimulation Decreases the Incidence of Amygdala‐kindled Seizures. Epilepsia. 46, 1-7. Hannesson, D., Mohapel, P., Corcoran, M., 2001. Dorsal hippocampal kindling selectively impairs spatial learning/short‐term memory. Hippocampus. 11, 275-286. Holmes, G.L., 1991. The long-term effects of seizures on the developing brain: clinical and laboratory issues. Brain and Development. 13, 393-409. Jahanshahi, A., Mirnajafi‐Zadeh, J., Javan, M., Mohammad‐Zadeh, M., Rohani, R., 2009. The antiepileptogenic effect of electrical stimulation at different low frequencies is accompanied with change in adenosine receptors gene expression in rats. Epilepsia. 50, 1768-1779. Kalynchuk, L., Fournier, N., 2009. Interictal anxiety in temporal lobe epilepsy. Encyclopedia of Basic Epilepsy Research. Vol., ed.^eds. San Diego: Elsevier Press/Academic Press. Kalynchuk, L.E., Wintink, A.J., 2005. A potential role for the hippocampus in the expression of kindling-induced fear. In Kindling 6. Vol., ed.^eds. Springer, pp. 285-294. 17
Kemp, A., Manahan-Vaughan, D., 2004. Hippocampal long-term depression and long-term potentiation encode different aspects of novelty acquisition. Proceedings of the National Academy of Sciences of the United States of America. 101, 8192-8197. Kemp, A., Manahan-Vaughan, D., 2007. Hippocampal long-term depression: master or minion in declarative memory processes? Trends in neurosciences. 30, 111-118. Krug, M., Koch, M., Grecksch, G., Schulzeck, K., 1997. Pentylenetetrazol kindling changes the ability to induce potentiation phenomena in the hippocampal CA1 region. Physiology & behavior. 62, 721-727. Larson, J., Xiao, P., Lynch, G., 1993. Reversal of LTP by theta frequency stimulation. Brain research. 600, 97-102. Leung, L.S., Boon, K.A., Kaibara, T., Innis, N.K., 1990. Radial maze performance following hippocampal kindling. Behavioural brain research. 40, 119-129. Leung, L.S., Shen, B., 1991. Hippocampal CA1 evoked response and radial 8-arm maze performance after hippocampal kindling. Brain research. 555, 353-357. Leung, L.S., Wu, C., 2003. Kindling suppresses primed-burst-induced long-term potentiation in hippocampal CA1. Neuroreport. 14, 211-214. Maru, E., Tatsuno, J., Okamoto, J., Ashida, H., 1982. Development and reduction of synaptic potentiation induced by perforant path kindling. Experimental neurology. 78, 409-424. McEachern, J.C., Shaw, C.A., 1996. An alternative to the LTP orthodoxy: a plasticity-pathology continuum model. Brain research reviews. 22, 51-92. McNaughton, B.L., Morris, R.G., 1987. Hippocampal synaptic enhancement and information storage within a distributed memory system. Trends in neurosciences. 10, 408-415. Mohammad-Zadeh, M., Mirnajafi-Zadeh, J., Fathollahi, Y., Javan, M., Ghorbani, P., Sadegh, M., Noorbakhsh, S.M., 2007. Effect of low frequency stimulation of perforant path on kindling rate and synaptic transmission in the dentate gyrus during kindling acquisition in rats. Epilepsy research. 75, 154-161. Mohammad-Zadeh, M., Mirnajafi-Zadeh, J., Fathollahi, Y., Javan, M., Jahanshahi, A., Noorbakhsh, S., Motamedi, F., 2009. The role of adenosine A 1 receptors in mediating the inhibitory effects of low frequency stimulation of perforant path on kindling acquisition in rats. Neuroscience. 158, 1632-1643. 18
Nabavi, S., Fox, R., Proulx, C.D., Lin, J.Y., Tsien, R.Y., Malinow, R., 2014. Engineering a memory with LTD and LTP. Nature. O'Dell, T.J., Kandel, E.R., 1994. Low-frequency stimulation erases LTP through an NMDA receptor-mediated activation of protein phosphatases. Learning & Memory. 1, 129-139. Palizvan, M., Fathollahi, Y., Semnanian, S., 2005. Epileptogenic insult causes a shift in the form of long-term potentiation expression. Neuroscience. 134, 415-423. Palizvan, M.R., Fathollahi, Y., Semnanian, S., Hajezadeh, S., Mirnajafizadh, J., 2001. Differential effects of pentylenetetrazol-kindling on long-term potentiation of population excitatory postsynaptic potentials and population spikes in the CA1 region of rat hippocampus. Brain research. 898, 82-90. Pastalkova, E., Serrano, P., Pinkhasova, D., Wallace, E., Fenton, A.A., Sacktor, T.C., 2006. Storage of spatial information by the maintenance mechanism of LTP. Science. 313, 1141-1144. Racine, R., Newberry, F., Burnham, W., 1975. Post-activation potentiation and the kindling phenomenon. Electroencephalography and clinical neurophysiology. 39, 261-271. Racine, R., 1978. Kindling: the first decade. Neurosurgery. 3, 234-252. Racine, R., Moore, K.-A., Evans, C., 1991. Kindling-induced potentiation in the piriform cortex. Brain research. 556, 218-225. Racine, R.J., 1972. Modification of seizure activity by electrical stimulation: II. Motor seizure. Electroencephalography and clinical neurophysiology. 32, 281-294. Robinson, G.B., Sclabassi, R.J., Berger, T.W., 1991. Kindling-induced potentiation of excitatory and inhibitory inputs to hippocampal dentate granule cells. I. Effects on linear and nonlinear response characteristics. Brain research. 562, 17-25. Rüthrich, H., Grecksch, G., Krug, M., 2001. Development of long-lasting potentiation effects in the dentate gyrus during pentylenetetrazol kindling. International Journal of Developmental Neuroscience. 19, 247-254. Sadegh, M., Mirnajafi-Zadeh, J., Javan, M., Fathollahi, Y., Mohammad-Zadeh, M., Jahanshahi, A., Noorbakhsh, S., 2007. The role of galanin receptors in anticonvulsant effects of lowfrequency stimulation in perforant path–kindled rats. Neuroscience. 150, 396-403.
19
Schiller, Y., Bankirer, Y., 2007. Cellular mechanisms underlying antiepileptic effects of low-and high-frequency electrical stimulation in acute epilepsy in neocortical brain slices in vitro. Journal of neurophysiology. 97, 1887-1902. Shahpari, M., Mirnajafi-Zadeh, J., Firoozabadi, S.M.P., Yadollahpour, A., 2012. Effect of lowfrequency electrical stimulation parameters on its anticonvulsant action during rapid perforant path kindling in rat. Epilepsy research. 99, 69-77. Soderling, T.R., Derkach, V.A., 2000. Postsynaptic protein phosphorylation and LTP. Trends in neurosciences. 23, 75-80. Sutherland, R.J., Leung, L., Weisend, M.P., Schlife, J., McDonald, R.J., 1997. An evaluation of the effect of partial hippocampal kindling on place navigation by rats in the Morris water task. Psychobiology. 25, 126-132. Sutula, T., Steward, O., 1986. Quantitative analysis of synaptic potentiation during kindling of the perforant path. Journal of neurophysiology. 56, 732-746. Teyler, T.J., Discenna, P., 1984. Long-term potentiation as a candidate mnemonic device. Brain research reviews. 7, 15-28. Teyler, T.J., DiScenna, P., 1985. The role of hippocampus in memory: a hypothesis. Neuroscience & Biobehavioral Reviews. 9, 377-389. Volcy, G.M., 2003. [Mesial temporal lobe epilepsy: its physiopathology, clinical characteristics, treatment and prognosis]. Revista de neurologia. 38, 663-667. Yamamoto, J., Ikeda, A., Satow, T., Takeshita, K., Takayama, M., Matsuhashi, M., Matsumoto, R., Ohara, S., Mikuni, N., Takahashi, J., 2002. Low‐frequency Electric Cortical Stimulation Has an Inhibitory Effect on Epileptic Focus in Mesial Temporal Lobe Epilepsy. Epilepsia. 43, 491-495. Yan, W.-W., Wang, C.-Y., Zeng, J., Liu, Q.-Y., Xu, S.-T., Liu, W.-X., Xiao, P., Li, C.-H., 2014. Low-frequency stimulation of dorsal norephinephrine bundle reverses behavioral longterm potentiation and learning performance in rats. Neuroscience. 265, 238-244. Zhao, D., Leung, L.S., 1991. Effects of hippocampal kindling on paired-pulse response in CA1 in vitro. Brain research. 564, 220-229. Zhao, D., Leung, L.S., 1992. Hippocampal kindling induced paired-pulse depression in the dentate gyrus and paired-pulse facilitation in CA3. Brain research. 582, 163-167. 20
21
Figures legends Fig. 1. Time-line diagram showing the protocol used for LFS application in animals of kindled+LFS group.
Fig. 2. Effect low-frequency stimulation (LFS) on kindling-induced impairment in LTP induction in CA1 area of the hippocampus at 48h after kindling stimulation. (A) schematic figure showing the place of stimulating and recording electrodes in hippocampal slice (DG: dentate gyrus; sch: Schaffer collaterals). (B) Sample records of field excitatory post-synaptic potentials (fEPSP) recording at 48 h after the last kindling stimulation in control, kindled (K-48 h), kindled+LFS (KLFS-48 h) and LFS (LFS-48 h) groups. Numbers 1 and 2 show the time of fEPSP recording according to the time-course graphs in C. (C) Time-course of fEPSP before and after application of primed-burst stimulation (PBS) in experimental groups. Data are plotted as the average percentage change from baseline responses. Each point shows the mean ± SEM. Application of PBS could not induce LTP in K-48 h group. However, application of LFS (in KLFS-48 h group) returned the ability of LTP induction. Application of LFS alone had also inhibitory effect on LTP induction. (D) Bar diagrams showing the changes in percentage of fEPSP slope after application of PBS (from 0 to 45 min) in different experimental groups. Each bar is mean ± SEM . *** p<0.001 compared to control group. # p<0.05 compared to K-48 h group (n=7 in all groups).
Fig. 3. Effect low-frequency stimulation (LFS) on kindling-induced impairment in LTP induction in CA1 area of the hippocampus at one week after kindling stimulation. (A) Sample records of field excitatory post-synaptic potentials (fEPSP) recording at one week after the last 22
kindling stimulation in control, kindled (K-1 week), kindled+LFS (KLFS-1 week) and LFS (LFS-1 week) groups. Numbers 1 and 2 show the time of fEPSP recording according to the timecourse graphs in B. (B) Time-course of fEPSP before and after application of primed-burst stimulation (PBS) in experimental groups. Data are plotted as the average percentage change from baseline responses. Each point shows the mean ± SEM. Application of PBS did not induce LTP in K-1 week group. However, application of LFS (in KLFS-1 week group) returned the ability of LTP induction. Application of LFS alone had also inhibitory effect on LTP induction. (C) Bar diagrams showing the changes in percentage of fEPSP slope after application of PBS (from 0 to 45 min) in different experimental groups. Each bar is mean ± SEM. *** p<0.001 compared to control group.
#
p<0.05 compared to K-1 week group (n=7 in all groups).
Fig. 4. Effect low-frequency stimulation (LFS) on kindling-induced impairment in LTP induction in CA1 area of the hippocampus at one month after kindling stimulation. (A) Sample records of field excitatory post-synaptic potentials (fEPSP) recording at one month after the last kindling stimulation in control, kindled (K-1 week), kindled+LFS (KLFS-1 week), kindled+2 LFS (K2LFS-1 month) and LFS (LFS-1 month) groups. Numbers 1 and 2 show the time of fEPSP recording according to the time-course graphs in B. (B) Time-course of fEPSP before and after application of primed-burst stimulation (PBS) in experimental groups. Data are plotted as the average percentage change from baseline responses. Each point shows the mean ± SEM. Application of PBS could not induce LTP in K-1 month and KLFS-1 month groups. However, application of additional LFS packages at one week after the first packages (see text) (K2LFS-1 month group) returned the ability of LTP induction in hippocampal slices prepared one month after the last kindling stimulation. Application of LFS alone had also inhibitory effect on LTP 23
induction. (C) Bar diagrams showing the changes in percentage of fEPSP slope after application of PBS (from 0 to 45 min) in different experimental groups. Each bar is mean ± SEM. *** p<0.001 compared to control group. ## p<0.01 compared to K-1 month group (n=7 in all groups).
Fig. 5. Input-output (I-O) curves were acquired according to the slope of field excitatory postsynaptic potential (fEPSP) in response to different stimulus intensities in the CA1 area of the hippocampus. I-O curves were drawn at 48 h (A), 1 week (B) and 1 month (C) after the last kindling stimulation in different experimental groups. There was a significant left-side shift in IO curve of kindled animals at 48 h (K-48 h group), 1 week (K-1 week) and 1 month (K-1 month) following the last kindling stimulation. Application of LFS in kindled animals completely returned this left-side shift toward control curve in hippocampal slices prepared at 48 h (KLFS48 h) and one week (KLFS- 1 week), but not one month (KLFS-1 month) after the last kindling stimulation. However, application of additional LFS packages at one week after the first packages (see text) could completely return the I-O curve toward control situation at one-month after kindling (K2LFS-1 month group).
24
Highlights: Long-term potentiation (LTP) cannot be induced in hippocampus up to 1 month after kindling. Low-frequency stimulation (LFS) alleviated kindling-induced deficit in LTP induction. Improving effect of LFS on LTP induction remained up to one week in kindled animals. LFS effects extended up to one month by increasing the number of its applications.
25
26
27
28
29
30