Major depressive disorder is the most common psychiatric disorder worldwide. To understand mechanisms and search for new approaches to treating depression, animal models are crucial. Chronic mild ...stress (CMS) is the most used animal model of depression. Although CMS is considered a robust model of depression, conflicting results have been reported for emotion-related behaviors, which the intrinsic characteristics of each rodent strain could explain. To further shed light on the impact of genetic background on the relevant parameters commonly addressed in depression, we examined the effect of 4-weeks CMS on anxiety and depression-related behaviors and body weight gain in three strain mice (BALB/c, C57BL/6, and CD1) of both sexes. CMS reduced body weight gain in C57BL/6NCrl and CD1 male mice. C57BL/6 animals exhibited a more pronounced anxious-like behavior than CD1 and BALB/c mice in the light-dark box (LDB) and the elevated plus maze (EPM) tests, whereas BALB/c animals exhibited the more robust depressive-like phenotype in the splash test (ST), tail suspension test (TST) and forced-swimming test (FST). Under CMS, exposure did not affect anxiety-related behaviors in any strain but induced depression-like behaviors strain-dependently. CMS C57BL/6 and CD1 mice of both sexes showed depression-like behaviors, and CMS BALB/c male mice exhibited reduced depressive behaviors in the FST. These results suggest a differential effect of stress, with the C57BL/6 strain being more vulnerable to stress than the CD1 and BALB/c strain mice. Furthermore, our findings emphasize the need for researchers to consider mouse strains and behavioral tests in their CMS experimental designs.
•Male and female C57BL/6 mice show a more robust basal anxious-like behavior.•Male and female BALB/c mice show a more robust basal depressive-like behavior.•CMS induces apathy-and-depressive-like behavior in male and female C57BL/6 mice.•CMS induces depressive-like behavior in male and female CD1 mice.•Under CMS, BALB/c mice show from negligible to inverse-expected behavior.
Animal models are crucial to understanding the mechanisms underlying the deleterious consequences of early-life stress. Here, we aimed to examine the effect of the limited bedding nesting (LBN) ...paradigm on early life development milestones and anxiety- and/or depression-like behavior in adolescent and adult mice from two inbred mice of both sexes. C57BL/6NCrl and BALB/c litters were exposed to the LBN paradigm postnatal day (PND) 2-9. Maternal behavior recording occurred on PND 3-9, and pups were weighed daily and examined to verify the eye-opening on PND 10-22. The male and female offspring underwent evaluation in the open field test, elevated plus-maze, and the forced swimming test during adolescence (PND 45-49) and adulthood (PND 75-79). We found that LBN impaired the maternal behavior patterns of both strain dams, mainly on C57BL/6NCrl strain. Also, LBN delayed the pup's eye-opening time and reduced body weight gain, impacting C57BL/6NCrl pups more. We also found that LBN decreased anxiety-related indices in adolescent and adult male but not female mice of both strains. Furthermore, LBN decreased depression-related indices only adolescent female and male BALB/c and female but not male C57BL/6NCrl mice. These findings reinforce the evidence that the LBN paradigm impairs the maternal behavior pattern and pup's early developmental milestones but does not induce anxiety- or depressive-like behavior outcomes during later life.
Abstract Small conductance calcium (Ca2+ ) activated SK channels are critical regulators of neuronal excitability in hippocampus. Accordingly, these channels are thought to play a key role in ...controlling neuronal activity in acute models of epilepsy. In this study, we investigate the expression and function of SK channels in the pilocarpine model of mesial temporal lobe epilepsy. For this purpose, protein expression was assessed using western blotting assays and gene expression was analyzed using TaqMan-based probes and the quantitative real-time polymerase chain reaction (qPCR) comparative method delta-delta cycle threshold (△△CT) in samples extracted from control and epileptic rats. In addition, the effect of SK channel antagonist UCL1684 and agonist NS309 on CA1 evoked population spikes was studied in hippocampal slices. Western blotting analysis showed a significant reduction in the expression of SK1 and SK2 channels at 10 days following status epilepticus (SE), but levels recovered at 1 month and at more than 2 months after SE. In contrast, a significant down-regulation of SK3 channels was detected after 10 days of SE. Analysis of gene expression by qPCR revealed a significant reduction of transcripts for SK2 ( Kcnn1 ) and SK3 ( Kcnn3 ) channels as early as 10 days following pilocarpine-induced SE and during the chronic phase of the pilocarpine model. Moreover, bath application of UCL1684 (100 nM for 15 min) induced a significant increase of the population spike amplitude and number of spikes in the hippocampal CA1 area of slices obtained control and chronic epileptic rats. This effect was obliterated by co-administration of UCL1684 with SK channel agonist NS309 (1 μM). Application of NS309 failed to modify population spikes in the CA1 area of slices taken from control and epileptic rats. These data indicate an abnormal expression of SK channels and a possible dysfunction of these channels in experimental MTLE.
Abstract In the hippocampus, BK channels are preferentially localized in presynaptic glutamatergic terminals including mossy fibers where they are thought to play an important role regulating ...excessive glutamate release during hyperactive states. Large conductance calcium-activated potassium channels (BK, MaxiK, Slo) have recently been implicated in the pathogenesis of genetic epilepsy. However, the role of BK channels in acquired mesial temporal lobe epilepsy (MTLE) remains unknown. Here we used immunohistochemistry, laser scanning confocal microscopy (LSCM), Western immunoblotting and RT-PCR to investigate the expression pattern of the α-pore-forming subunit of BK channels in the hippocampus and cortex of chronically epileptic rats obtained by the pilocarpine model of MTLE. All epileptic rats experiencing recurrent spontaneous seizures exhibited a significant down-regulation of BK channel immunostaining in the mossy fibers at the hilus and stratum lucidum of the CA3 area. Quantitative analysis of immunofluorescence signals by LSCM revealed a significant 47% reduction in BK channel immunofluorescent signals in epileptic rats when compared to age-matched non-epileptic control rats. These data correlate with a similar reduction in BK channel protein levels and transcripts in the cortex and hippocampus. Our data indicate a seizure-related down-regulation of BK channels in chronically epileptic rats. Further functional assays are necessary to determine whether altered BK channel expression is an acquired channelopathy or a compensatory mechanism affecting the network excitability in MTLE. Moreover, seizure-mediated BK down-regulation may disturb neuronal excitability and presynaptic control at glutamatergic terminals triggering exaggerated glutamate release and seizures.
Abstract Group II metabotropic (mGlu II) receptor subtypes mGlu2 and mGlu3 are important modulators of synaptic plasticity and glutamate release in the brain. Accordingly, several pharmacological ...ligands have been designed to target these receptors for the treatment of neurological disorders characterized by anomalous glutamate regulation including epilepsy. In this study, we examine whether the expression level and function of mGlu2 and mGlu3 are altered in experimental epilepsy by using immunohistochemistry, Western blot analysis, RT-PCR and extracellular recordings. A down-regulation of mGlu2/3 protein expression at the mossy fiber pathway was associated with a significant reduction in mGlu2/3 protein expression in the hippocampus and cortex of chronically epileptic rats. Moreover, a reduction in mGlu2 and mGlu3 transcripts levels was noticed as early as 24 h after pilocarpine-induced status epilepticus (SE) and persisted during subsequent “latent” and chronic periods. In addition, a significant impairment of mGlu II-mediated depression of field excitatory postsynaptic potentials at mossy fiber-CA3 synapses was detected in chronically epileptic rats. Application of mGlu II agonists (2S,2′R,3′R)-2-(2′,3′-dicarboxycyclopropyl)glycine (DCG-IV) induced a significant reduction of the fEPSP amplitude in control rats, but not in chronic epileptic rats. These data indicate a long-lasting impairment of mGlu2/3 expression that may contribute to abnormal presynaptic plasticity, exaggerate glutamate release and hyperexcitability in temporal lobe epilepsy.
Abstract Group II metabotropic glutamate (mGlu II) receptors subtype 2 and 3 (mGlu2 and mGlu3) are subtle regulators of neuronal excitability and synaptic plasticity in the hippocampus. In recent ...years, researchers have investigated the potential neuroprotective and anticonvulsant effects of compounds acting on mGlu II receptors. However, abnormal expression and function of mGlu2 and mGlu3 have been reported in temporal lobe epilepsy, a phenomena that may limit the therapeutic effectiveness of these potentially new antiepileptic drugs. Here, we investigated seizure-induced changes in mGlu2 and mGlu3 mRNA following pilocarpine-inducted status epilepticus (SE) and subsequent epileptogenesis. Relative changes in gene expression were assessed by comparative analysis of quantitative real-time PCR (qrtPCR) by the delta–delta CT method. Pilocarpine-treated and control rats were sacrificed at different periods (24 h, 10 days, one month and more than two months) following SE. Total RNA was isolated from microdissected dentate gyrus and processed for RT-PCR and qrtPCR using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as an endogenous control gene. Analysis of relative quantification (RQ) ratios of mGlu2 and mGlu3 mRNA expression revealed a significant down-regulation of both targets at 24 h after SE. Gene expression partially recovered at 10 days following SE reaching control levels at one month after SE. Two month after SE, mGlu2 mRNA expression was significantly down-regulated to ~ 41% of control expression whereas mGlu3 mRNA was comparable to control levels. Our data indicate that mGlu2 and mGlu3 expression is dynamically down-regulated or selectively enhanced during critical periods of epileptogenesis. Seizure-induced differential dysregulation of mGlu2 and mGlu3 receptors may affect the availability of these molecular targets for therapeutic compounds in epilepsy.
Functional properties of large conductance Ca(2+) activated potassium (BK) channels are determined by complex alternative splicing of the Kcnma1 gene encoding the alpha pore-forming subunit. ...Inclusion of the STREX exon in a C-terminal splice site is dynamically regulated and confers enhanced Ca(2+) sensitivity and channel inhibition via cAMP-dependent phosphorylation. Here, we describe a real time quantitative PCR (qPCR) approach to investigate relative changes in the expression of STREX and ZERO splice variants using a newly designed set of probes and primers for TaqMan-based qPCR analysis of cDNA from the rat dentate gyrus at different time points following pilocarpine-induced status epilepticus. Reduction in Kcnma1 gene expression is associated with a relative increase of STREX splice variant. Relative expression of STREX variant mRNA was increased at 10 days and at more than 1 month following status epilepticus. The biological consequences of seizure-related changes in alternative splicing of Kcnma1 deserve additional investigation.
Epilepsy is characterized by hyperexcitability of hippocampal networks, excessive release of glutamate, and progressive neurodegeneration. Presynaptic group II metabotropic receptors (mGluR2 and ...mGluR3) are among different mechanisms that modulate presynaptic release of glutamate, especially at the mossy fibers in the hippocampus. Here, we explore whether mGluR2/3 expression is affected in a rat model of temporal lobe epilepsy obtained via pilocarpine-induced
status epilepticus (SE). Immunohistochemical assays were performed in age-matched controls and two groups of epileptic rats sacrificed at 25–35 days (1 month post-SE) and at 55–65 days (2 months post-SE) following SE onset. A dramatic lessening of mGluR2/3 immunofluorescence was observed at CA1 and CA3
stratum lacunosum/molecular (SLM) declining to 60% and 68% of control values in 1-month and 2-month post-SE, respectively. Additionally, thickness of mGluR2/3-stained SLM layer narrowed up to 70% of controls indicating atrophy at this branch of the perforant path. Epileptic rats exhibited a marked and progressive down-regulation of mGluR2/3 expression in mossy fiber at
hilus and CA3
stratum lucidum in contrast with an enhanced expression of vesicular glutamate transporter type 1 (VGluT1) at the mossy fibers. Intense VGluT1 punctated staining was detected at the inner third molecular layer indicating glutamatergic sprouting. In the molecular layer, mGluR2/3 labeling slightly declined in the 1-month post-SE group but then increased in the 2-month post-SE group although it was diffusely distributed. Down-regulation of mGluR2/3 at the mossy fibers and the SLM may render epileptic hippocampal networks hyperexcitable and susceptible to glutamate-mediated excitotoxicity and neurodegeneration.
Abstract Alzheimer's disease (AD) is a progressive, neurodegenerative brain disorder characterized by extracellular accumulations of amyloid β (Aβ) peptides, intracellular accumulation of abnormal ...proteins, and early loss of basal forebrain neurons. Recent studies have indicated that the conformation of Aβ is crucial for neuronal toxicity, with intermediate misfolded forms such as oligomers being more toxic than the final fibrillar forms. Our previous work shows that Aβ blocks the potassium (K+ ) currents IM and IA in septal neurons, increasing firing rates, diminishing rhythmicity and firing coherence. Evidence also suggests that oxidative stress (OS) plays a role in AD pathogenesis. Thus we wished to determine the effect of oligomeric and fibrillar forms of Aβ1–42 on septohippocampal damage, oxidative damage, and dysfunction in AD. Oligomeric and fibrillar forms of Aβ1–42 were injected into the CA1 region of the hippocampus in live rats. The rats were sacrificed 24 hours and 1 month after Aβ or sham injection to additionally evaluate the temporal effects. The expression levels of the K+ voltage-gated channel, KQT-like subfamily, member 2 (KCNQ2 ) and the OS-related genes superoxide dismutase 1, 8-oxoguanine DNA glycosylase, and monamine oxidase A, were analyzed in the hippocampus, medial, and lateral septum. Our results show that both forms of Aβ exhibit time-dependent differential modulation of OS and K+ channel genes in the analyzed regions. Importantly, we demonstrate that Aβ injected into the hippocampus triggered changes in gene expression in anatomical regions distant from the injection site. Thus the Aβ effect was transmitted to anatomically separate sites, because of the functional coupling of the brain structures.
Abstract Voltage gated K+ channels (Kv) are a highly diverse group of channels critical in determining neuronal excitability. Deficits of Kv channel subunit expression and function have been ...implicated in the pathogenesis of epilepsy. In this study, we investigate whether the expression of the specific subunit Kv3.4 is affected during epileptogenesis following pilocarpine-induced status epilepticus . For this purpose, we used immunohistochemistry, Western blotting assays and comparative analysis of gene expression using TaqMan-based probes and delta-delta cycle threshold (ΔΔCT) method of quantitative real-time polymerase chain reaction (qPCR) technique in samples obtained from age-matched control and epileptic rats. A marked down-regulation of Kv3.4 immunoreactivity was detected in the stratum lucidum and hilus of dentate gyrus in areas corresponding to the mossy fiber system of chronically epileptic rats. Correspondingly, a 20% reduction of Kv3.4 protein levels was detected in the hippocampus of chronic epileptic rats. Real-time quantitative PCR analysis of gene expression revealed that a significant 33% reduction of transcripts for Kv3.4 (gene Kcnc4 ) occurred after 1 month of pilocarpine-induced status epilepticus and persisted during the chronic phase of the model. These data indicate a reduced expression of Kv3.4 channels at protein and transcript levels in the epileptic hippocampus. Down-regulation of Kv3.4 in mossy fibers may contribute to enhanced presynaptic excitability leading to recurrent seizures in the pilocarpine model of temporal lobe epilepsy.