Hypothalamic KiSS1/GPR54 gene expressions and luteinizing hormone plasma secretion in morphine treated male rats

Background: The inhibitory effects of morphine and the stimulatory influence of kisspeptin signaling have been demonstrated on gonadotropin releasing hormone [GnRH]/luteinizing hormone [LH] release. Hypothalamic kisspeptin is involved in relaying the environmental and metabolic information to reproductive axis. In the present study, the role of kisspeptin/ GPR54 signaling system was investigated on relaying the inhibitory influences of morphine on LH hormone secretion

Materials and Methods: In this experimental study, 55 wistar male rats weighing 230-250 g were sub-grouped in 11 groups [in each group n=5] receiving saline, kisspeptin [1 nmol], peptide234 [P234, 1 nmol], morphine [5 mg/kg], naloxone [2 mg/kg], kisspeptin/P234, morphine/naloxone, kisspeptin/morphine, kisspeptin/naloxone, P234/morphine or P234/naloxone respectively. Blood samples were collected via tail vein. Mean plasma [LH] concentrations and mean relative KiSS1 or GPR54 mRNA levels were determined by radioimmunoassay [RIA] and real time reverse transcriptase-polymerase chain reaction [RT-PCR], respectivwely

Results: Morphine significantly decreased mean plasma LH concentration and mean relative KiSS1 gene expression compared to saline; while it did not significantly decrease mean relative GPR54 gene expression compared to saline. Naloxone significant increased mean LH level and mean relative KiSS1 gene expression compared to saline; while it did not significantly increase mean relative GPR54 gene expression compared to saline. Injections of kisspeptin plus morphine significantly increased mean LH concentration compared to saline or morphine, while simultaneous infusions of them significantly declined mean plasma LH level compared to kisspeptin. In kisspeptin/naloxone group mean plasma LH level was significantly increased compared to saline or naloxone. Co-administration of P234/morphine significantly decreased mean LH concentration compared to saline

Conclusion: Down regulation of KiSS1 gene expression may be partly involved in the mediating the inhibitory effects of morphine on reproductive axis

Impaired memory and evidence of histopathology in CA1 pyramidal neurons through Injection of Abeta1-42 peptides into the frontal cortices of rat

Introduction: Alzheimer's disease [AD] is one of the most common neurodegenerative disorders, which has much benefited from animal models to find the basics of its pathophysiology. In our previous work [Haghani, Shabani, Javan, Motamedi, and Janahmadi, 2012], a non-transgenic rat model of AD was used in electrophysiological studies. However, we did not investigate the histological aspects in the mentioned study

Methods: An AD model was developed through bilateral injection of amyloid-beta peptides [Abeta] into the frontal cortices. Behavioral and histological methods were used to assess alterations in the memory and [ultra]structures. Furthermore, melatonin has been administered to assess its efficacy on this AD model

Results: Passive avoidance showed a progressive decline in the memory following Abeta injection. Furthermore, Nissl staining showed that Abeta neurotoxicity caused shrinkage of the CA1 pyramidal neurons. Neurodegeneration was clearly evident from Fluoro-jade labeled neurons in Abeta treated rats. Moreover, higher NF-kappaB immunoreactive CA1 pyramidal neurons were remarkably observed in Abeta treated rats. Ultrastructural analysis using electron microscopy also showed the evidence of subcellular abnormalities. Melatonin treatment in this model of AD prevented Abeta- induced increased NF-kappaB from immunoreaction and neurodegeneration

Discussion: This study suggests that injection of Abeta into the frontal cortices results in the memory decline and histochemical disturbances in CA1 pyramidal neurons. Furthermore, melatonin can prevent several histological changes induced by Abeta

Suppressive effects of resveratrol treatment on the intrinsic evoked excitability of CA1 pyramidal neurons

Objective: Resveratrol, a phytoalexin, has a wide range of desirable biological actions. Despite a growing body of evidence indicating that resveratrol induces changes in neu-ronal function, little effort, if any, has been made to investigate the cellular effect of resveratrol treatment on intrinsic neuronal properties

Materials and Methods: This experimental study was performed to examine the acute effects of resveratrol [100 NM] on the intrinsic evoked responses of rat Cornu Ammonis [CA1] pyramidal neurons in brain slices, using whole cell patch clamp recording under current clamp conditions

Results: Findings showed that resveratrol treatment caused dramatic changes in evoked responses of pyramidal neurons. Its treatment induced a significant [P<0.05] increase in the after hyperpolarization amplitude of the first evoked action potential. Resveratrol-treated cells displayed a significantly broader action potential [AP] when compared with either control or vehicle-treated groups. In addition, the mean instantaneous firing frequency between the first two action potentials was significantly lower in resveratrol-treated neurons. It also caused a significant reduction in the time to maximum decay of AP. The rheobase current and the utilization time were both significantly greater following resveratrol treatment. Neurons exhibited a significantly depolarized voltage threshold when exposed to resveratrol

Conclusion: Results provide direct electrophysiological evidence for the inhibitory effects of resveratrol on pyramidal neurons, at least in part, by reducing the evoked neural activity

Anethole, a medicinal plant compound, decreases the production of pro-inflammatory TNF-alpha and IL[-1]beta in a rat model of LPS-induced periodontitis

Periodontitis [PD] is known to be one of most prevalent worldwide chronic inflammatory diseases. There are several treatments including antibiotics for PD; however, since drug resistance is an increasing problem, new drugs particularly derived from plants with fewer side effects are required. The effects of trans-anethole on IL[-1] beta and TNF-alpha level in a rat model of PD were investigated and compared to ketoprofen. Eschericia coli lipopolysaccharide [LPS, 30 microg] was injected bilaterally into the palatal gingiva [3 microL/site] between the upper first and second molars every two days for 10 days in anesthetized rats. Administration of either trans-anethole [10 or 50 mg/Kg, i.p.] or ketoprofen [10 mg/Kg, i.p.] was started 20 minute before LPS injection and continued for 10 days. Then, IL[-1]beta and TNF-alpha levels were measured in blood samples by ELISA at day 0 [control] and at day 10. Anethole at both concentrations significantly suppressed IL[-1]beta and TNF-alpha production when compared to LPS-treated rats. The suppressive effects of anethole on LPS-induced pro-inflammatory cytokines were almost similar as seen with ketoprofen. In conclusion, the present results suggest that anethole may have a potent inhibitory effect on PD through suppression of pro-inflammatory molecules; therefore it could be a novel therapeutic strategy for PD

effect of paxilline on early alterations of electrophysiological properties of dentate gyrus granule cells in pilocarpine-treated rats

The dentate gyrus of hippocampus has long been considered as a focal point for studies on mechanisms responsible for the development of temporal lobe epilepsy [TLE]. Change in intrinsic properties of dentate gyrus granule cells [GCs] has been considered as an important factor responsible in temporal lobe seizures. In this study, we evaluated the intrinsic properties of GCs, during acute phase of seizure [24 h after i.p. injection of pilocarpine] compared to sham group using whole cell patch-clamp recordings. Our results showed a significant increase in the number of action potentials [APs] after applying depolarizing currents of 200 pA [p < 0.01] and 250pA [p < 0.05] compared to sham group. The evaluation of AP properties revealed a decrease in half-width of AP in GCs of seizure group [1.27 +/- 0.03 ms] compared to sham group [1.60 +/- 0.11]. Moreover, addition of BAPTA to pipette solution prevented changes in AP half-width in seizure group [1.71 +/- 0.11 ms] compared to sham group [1.91 +/- 0.08 ms]. In contrast, an increase in the amplitude of fast afterhyperpolarization was observed in GCs of seizure group [-11.68 +/- 0.72 mV] compared to sham group [-8.28 +/- 0.59 mV]. Also, GCs of seizure group showed a significant increase in both firing rate and instantaneous firing frequency at depolarizing currents of 200 pA [P < 0.01] and 250 pA [P < 0.05] compared to sham group. The changes in electrophysiological properties of GCs were attenuated after bath application of paxilline suggesting possible involvement of large conductance Ca[2+]- activated K[+] channel [BK channel]. Our results suggested the possible involvement of certain potassium channels in early changes of intrinsic properties of GCs which eventually facilitate TLE development

comparison of the effects of acute and repeated morphine administration on fast synaptic transmission in magnocellular neurons of supraoptic nucleus, plasma vasopressin levels, and urine volume of male rats

The activity of the magnocellular neurons [MCNs] of supraoptic nucleus [SON] is regulated by a variety of excitatory and inhibitory inputs. Opioids are one of the important compounds that affect these inputs at SON synapses. In this study, whole-cell patch clamp recording of SON neurons was used to investigate the effect of acute and repeated morphine administration on spontaneous inhibitory and excitatory post synaptic currents [sIPSCs and sEPSCs] in MCNs. While acute bath application of morphine to brain slice of intact rat produced an increase in sEPSCs frequency and a decrease in sIPSCs frequency, repeated in vivo administration of morphine produced opposite effect. Moreover, repetitive i.c.v. administration of morphine for three consecutive days caused significant increase in urine volume, but had no significant alteration in water consumption compared to control group. The increase in urine volume was consistent with a significant decrease in plasma arginine vasopressin [AVP] levels after repetitive i.p. morphine administration. The results suggest that acute administration of morphine stimulates whereas repeated administration of morphine inhibits the MCNs. Morphine-induced MCN inhibition could result in diminished plasma AVP levels and eventually an increase in urine volume of rats

Morphine attenuates testosterone response to central injection of kisspeptin in male rats

Background: Kisspeptin and naloxone stimulate the reproductive axis while morphine inhibits its function. We have investigated the effect of central injection of kisspeptin-10 on mean plasma testosterone concentration in morphine or naloxone pretreated rats

Materials and Methods: In this experimental study, 60 male Wistar rats that were divided into 12 groups [n=5 per group] received saline, kisspeptin [1 nmol, ICV], naloxone [2 mg/kg, subcutaneously], morphine [5 or 10 mg/kg, sc] or co-administrations of kisspeptin, morphine and naloxone at 09:00 - 09:30. In the co-administrated groups, kisspeptin was injected 15 minutes following morphine or naloxone injections. Blood samples were collected 60 minutes following injections via the tail vein. Plasma testosterone concentration was measured by a rat testosterone ELISA kit

Results: Central injection of kisspeptin or subcutaneous injection of naloxone significantly increased the mean plasma testosterone concentration compared to saline while subcutaneous injections of different doses of morphine [5 or 10 mg/kg] significantly decreased testosterone compared to saline. The results revealed that morphine significantly attenuated the testosterone increase after kisspeptin injection compared to kisspeptin while a stimulatory additive effect was observed in the kisspeptin/naloxone group compared to either naloxone or kisspeptin

Conclusion: Morphine and kisspeptin systems may interact with each other to control the hypothalamic-pituitary-gonadal [HPG] axis

rare case of klinefelter syndrome patient with quintuple mosaic karyotype, diagnosed by GTG-banding and FISH

Klinefelter syndrome [KS] is the most common sex chromosomal disorder in men. Most of these patients show the 47, XXY karyotype, whereas approximately 15% of them are mosaics with variable phenotype. A 39-year-old male investigated for primary infertility, was clinically normal with small firm testes and elevated levels of FSH, LH and low level of testosterone. Total azoospermia was confirmed on semen analysis. Testicular histopathology revealed no spermatogenesis and absence of germ cells. Karyotype from whole blood culture showed cells with 47, XXY/46, XX/ 45, X/48, XXXY/ 46, XY mosaicism. The predominant cell line was 47, XXY [83.67%]. This was confirmed by fluorescence in situ hybridization [FISH]. Also the presence of a small population of cells with the 48, XXXY and 45, X karyotypes was detected by FISH. This case illustrates the utility of FISH as an adjunct to conventional cytogenetics in assess the chromosome copy number in each cell line of a mosaic

Antibiotic supplements affect electrophysiological properties and excitability of rat hippocampal pyramidal neurons in primary culture

Introduction: antibiotic supplements are regularly used in neuronal culture media to control contamination; however, they can interfere with the neuronal excitability and affect electrophysiological properties. Therefore, in this study, the effect of penicillin/streptomycin supplements on the spontaneous electrophysiological activity of hippocampal pyramidal neurons was examined

Methods: electrophysiological whole-cell patch-clamp recordings from rat hippocampal pyramidal cells in primary culture were performed to investigate the effects of antibiotic supplements on the intrinsic excitability of cultured cells

Results: the present findings indicated that presence of antibiotic supplements [penicillin/streptomycin] in the culture medium altered the intrinsic electrical activity of hippocampal pyramidal neurons in primary culture. These alterations included: 1] depolarized resting membrane potential; 2] a significant enhancement in the after-hyperpolarization amplitude; 3] a significant increase in the area under the action potential and in the decay and rise time of the action potential; 4] a significant broadening of action potential and 5] a significant reduction in the firing frequency

Conclusion: these findings suggest that addition of antibiotic supplements to culture media influences the neuronal excitability and alters the electrophysiologicalproperties of cultured neurons, possibly through changing the ionic conductance underlying neuronal excitability. Iran. Biomed. J. 17 [2]: 101-106, 2013

Computational insights into the neuroprotective action of riluzole on 3-acetylpyridine-induced ataxia in rats

Intra-peritoneal administration of riluzole has been shown to preserve the membrane properties and firing characteristics of Purkinje neurons in a rat model of cerebellar ataxia induced by 3-acetylpyridine [3-AP]. However, the exact mechanism[s] by which riluzole restores the normal electrophysiological properties of Purkinje neurons is not completely understood. Changes in the conductance of several ion channels, including the BK channels, have been proposed as a neuro protective target of riluzole. In this study, the possible cellular effects of riluzole on Purkinje cells from 3-AP-induced ataxic rats that could be responsible for its neuro protective action have been investigated by computer simulations. This is a computational stimulation study. The simulation environment enabled a change in the properties of the specific ion channels as the possible mechanism of action of riluzole. This allowed us to study the resulted changes in the firing activity of Purkinje cells without concerns about its other effects and interfering parameters in the experiments. Simulations were performed in the NEURON environment [Version 7.1] in a time step of 25 micro s; analyses were conducted using MATLAB r2010a [The Mathworks]. Data were given as mean +/- SEM. Statistical analyses were performed by the student's t test, and differences were considered significant if p<0.05. The computational findings demonstrated that modulation of an individual ion channel current, as suggested by previous experimental studies, should not be considered as the only possible target for the neuro protective effects of riluzole to restore the normal firing activity of Purkinje cells from ataxic rats. Changes in the conductance of several potassium channels, including voltage-gated potassium [Kv1, Kv4] and big Ca[2+]-activated K[+] [BK] channels may be responsible for the neuro protective effect of riluzole against 3-AP induced alterations in the firing properties of Purkinje cells in a rat model of ataxia

Comparing the effects of mineral trioxide aggregate and calcium enriched mixture on neuronal cells using an electrophysiological approach

The main goal of this ex vivo study was to assess and compare the cellular and electrophysiological effects of two dental biomaterials, white mineral trioxide aggregate [WMTA] and calcium enriched mixture [CEM] cement, on neuronal cell excitability and electrical properties. A conventional intracellular current clamp technique was used to study the cellular effects of WMTA and CEM on the excitability, firing and the shape of action potential of neuronal soma membrane of F1 nerve cells. The dental biomaterials were prepared according to the manufacturers' directions and were applied to the bathing media and 0.05 mL of total mixture of each dental material at a distance of 3 mm from the cells. Findings indicated that exposure to both dental biomaterials shifted the irregular high frequency firing type observed in control conditions to a more regular low frequency firing pattern. Neuronal exposure to WMTA, but not CEM, significantly hyperpolarized the cell resting membrane potential. Both treatments significantly influenced the duration and the amplitude of action potentials. Extracellular application of either CEM or WMTA caused a significant increase in the after hyperpolarization [AHP] amplitude and AHP area, but the potentiating effect of WMTA was more effective than CEM. Treatment with WMTA or CEM resulted in a profound alteration in the firing behaviour of F1 cells and changed the AP characteristics. Both dental biomaterials reduced the neuronal activity possibly through enhancement of K[+] outward current. This may possibly explain the positive mechanisms of these biomaterials in regenerative endodontics, though further research is needed for such a conclusion

electrophysiological consequences of artemisia dracunculus L. [Tarragon] extract on pentylenetetrazomnduced epilepti-form activity in snail neurons

Plant extracts are of considerable interest because of their antiepileptic activities. However, the mechanisms of action are not clearly defined. Here, the effects of Artemisia dracunculus L [tarragon] leaves extract on excitability and electrophysiological characteristics of snail neurones were investigated, using an intracellular recording technique. Application of tarragon extract [0.05%] resulted in complete disappearance of paroxysmal depolarization shift [PDS] as elicited by pentylenetetrazol [PTZ], an epileptogenic drug. It also significantly decreased the firing frequency and shifted the firing pattern from bursting in the presence of PTZ to an irregular doublet activity. Changes in excitability properties were associated with a significant increase and decrease in the duration of action potential, and in the amplitude of after-hyperpolarization [AHP], respectively. When tarragon extract was applied alone, spontaneous activity became irregular and was interrupted by large inhibitory postsynaptic potentials [IPSPs], which disappeared following application of picrotoxin [100 micro M]. Tarragon also caused a significant decrease both in the amplitude of action potentials and AHP, and broadened the action potentials. However, pretreatment with extract did not prevent the induction of epileptiform activity by PTZ. The findings suggest that tarragon extract may affect membrane ion channels and/or GABAA receptors leading to a reduction in neuronal excitability

comparative study on anti-hyperalgesia effect of MTA and ketoprofen in inflammatory pain

Mineral trioxide aggregate [MTA] is an endodontic material with different clinical applications e.g. root-end filling, pulp capping and perforation repair. It has been reported to possess antimicrobial and antifungal activities. The aim of this study was to examine the effect of White MTA on formalin-induced hyperalgesia in a rat with inflammatory pain. Inflammatory pain was induced by subcutaneous [SC] injection of formalin [40 microL, 2.5%] into the rat upper lip. The nociceptive behavioral responses i.e. shaking of the lower jaw and face rubbing were quantified. 40 uL of eugenol [50 mg/kg], WMTA [20 mg/0.2 mL] or ketoprofen were injected solely or in combination with formalin 2.5% and the behavioral responses were compared with those observed after formalin treatment alone. One-way ANOVA, Tukey were used for analysis of data. Formalin 2.5% provoked a biphasic nociceptive response, with an early and short lasting first tonic phase followed by a second phase. Solely SC injection of either WMTA or ketoprofen [a non steroidal anti-inflammatory drug] did not stimulate any significant nociceptive behaviour. However, injection of eugenol [a pain relieving agent] induced the early phase not the tonic phase of nociceptive response. WMTA, eugenol or ketoprofen injection 20 min before formalin injection attenuated the first phase but somehow prevented the induction of the second phase of nociceptive responses which were produced by formalin. Behavioural nociceptive responses including shaking of the lower jaw and face rubbing were significantly reduced when the subject was pretreated with either WMTA or ketoprofen [P<0.001]. In this study, WMTA induced pain reduction by suppression of the formalin-induced nociceptive response

Modification of nifedipine inhibitory effect on calcium spike and L - Type calcium current by ethanol in F1 neuron of helix aspersa

There is strong evidence demonstrating that nifedipine dissolved in ethanol selectively inhibits only L-type Ca2+ current. In addition, acute ethanol exposure reduces voltage-dependent calcium currents. In the present study, we investigated the antagonistic effect of fixed concentration of nifedipine dissolved in different concentration of ethanol on L-type Ca2+ current. In a Na +/- K+ free solution, nifedipine dissolved in 60 and 120 mM ethanol decreased resting membrane potential of Ca2+ spikes and caused a significant reduction in amplitude, duration and an increase in threshold of Ca2+ spikes. Furthermore, Ca2+ current was inhibited by ethanol in a concentration-dependent manner, so that the reduction of L-type Ca2+ current by nifedipine/60 and 120 mM ethanol was statistically significant. Meanwhile, ethanol concentration-dependent response of Ca2+ currents was observed at its late component in more positive potentials. These results may be consistent with ethanol-dependent inhibition of L-type Ca2+ currents and ethanol-dependent enhancment of a Ca2 +/- activated potassium current