Serotonin-mediated modulation of Na+/K+ pump current in rat hippocampal CA1 pyramidal neurons
© Zhang et al; licensee BioMed Central Ltd. 2012
Received: 14 March 2010
Accepted: 19 January 2012
Published: 19 January 2012
The aim of this study was to investigate whether serotonin (5-hydroxytryptamine, 5-HT) can modulate Na+/K+ pump in rat hippocampal CA1 pyramidal neurons.
5-HT (0.1, 1 mM) showed Na+/K+ pump current (Ip) densities of 0.40 ± 0.04, 0.34 ± 0.03 pA/pF contrast to 0.63 ± 0.04 pA/pF of the control of 0.5 mM strophanthidin (Str), demonstrating 5-HT-induced inhibition of Ip in a dose-dependent manner in hippocampal CA1 pyramidal neurons. The effect was partly attenuated by ondasetron, a 5-HT3 receptor (5-HT3R) antagonist, not by WAY100635, a 5-HT1AR antagonist, while 1-(3-Chlorophenyl) biguanide hydrochloride (m-CPBG), a 5-HT3R specific agonist, mimicked the effect of 5-HT on Ip.
5-HT inhibits neuronal Na+/K+ pump activity via 5-HT3R in rat hippocampal CA1 pyramidal neurons. This discloses novel mechanisms for the function of 5-HT in learning and memory, which may be a useful target to benefit these patients with cognitive disorder.
5-HT, as a neurotransmitter or neuromodulator in the central nervous system, plays a critical role in the control of blood pressure, body temperature, sleep, depression, anxiety, epilepsy [1–4]. Additionally, the modulation of the serotonergic system affects long-term potentiation (LTP) and long-term depression (LTD), the likely neurophysiologic derivates of learning and memory formation, which has been involved in the treatment of Alzheimer's disease [5–8]. Some studies demonstrate that 5-HT1AR-knockout animals show a deficit in hippocampal-dependent learning and memory, such as the hidden platform (spatial) version of the Morris water maze and the delayed version of the Y maze , while the stimulation of 5-HT1AR mediates enhancement of LTP  and prevents the impairment of learning and memory [11, 12]. Therefore, the stimulation of 5-HT1AR may be useful in the symptomatic treatment of human memory disturbances. However, accumulated clinical reports support that the injection of 5-HT3R antagonists facilitates the induction of LTP, and enhances the retention and consolidation of memory in hippocampal dependent tasks [13–15]. These clinical application of 5-HT3R antagonists have been found to improve memory in schizophrenic or Alzheimer demented patients [16, 17]. Therefore, 5-HT3R also plays a critical role in cognitive function.
In addition to increasing neuronal excitability , inhibition of Na+/K+ pump activity can induce LTD whereas depotentiate LTP , and then cause impairment of learning and memory and amnesia [20, 21]. Herein, in the present study, we investigate if a relationship occurs between 5-HT and Na+/K+ pump in hippocampal CA1 pyramidal neurons, which may provide new insights in the mechanisms responsible for the 5-HT-mediated modulation of learning and memory.
Results and discussion
5-HT-mediated inhibition of Ip in rat hippocampal CA1 pyramidal neurons
0.5 mM Str often did not recover completely in hippocampal CA1 slices even after prolonged washout in the present study, consistent with the previous study that Na+/K+ pump inhibition by Str was effectively irreversible . Furthermore, 10 μM Str did not recover completely in rat ventral midbrain slices . Therefore, Str perfusion was applied one time in one brain slice.
Furthermore, some studies reported that the application of 1 μM 5-HT prevented depotentiation but not LTP induced by high-frequency stimulation, whereas bath application of 100 μM 5-HT blocked the induction of tetanus-induced LTP , consistent with the previous study that 5-HT (30 μM) prevented LTP induced by a primed burst in rat hippocampal CA1 region . Accordingly, different concentrations of 5-HT may have different modulation of learning and memory. Moreover, the inhibition of Na+/K+ pump activity can induce LTD whereas depotentiate LTP , and then cause impairment of learning and memory and amnesia [20, 21]. Herein, in the present study, 5-HT-mediated inhibition of Na+/K+ pump activity may disclose novel mechanisms in learning and memory. Further studies should be done to explore the mechanism.
5-HT mediated inhibiton of Ip via 5-HT3R not 5-HT1AR
In the presence of ondasetron, a 5-HT3R antagonist, 5-HT-mediated inhibition of Ip was blocked from 0.40 ± 0.04 to 0.61 ± 0.04 pA/pF (Figure 2), while m-CPBG, a 5-HT3R specific agonist, mimicked the effect of 5-HT on Ip (Figure 2). These results show, for the first time, that the inhibition of 5-HT-mediated Ip is primarily mediated by 5-HT3R in hippocampal CA1 pyramidal neurons. On the subcellular level, both presynaptic and postsynaptic 5-HT3R can be found. Presynaptic 5-HT3R is involved in mediating or modulating neurotransmitter release. Postsynaptic 5-HT3R is preferentially expressed on interneurons [36, 37], and there is also 5-HT3R in postsynaptic pyramidal neurons [38–42]. For example, electrophysiological studies in postsynaptic pyramidal neurons in the hippocampal CAl region or hippocampal primary cultures showed the activation of 5-HT3R [38–42]. Furthermore, 5-HT3R is a unique serotonin receptor as it acts as a ligand-gated ion channel, whereas all the other types of serotonin receptors belong to the G protein-coupled receptor superfamily, which may be the reason of 5-HT3R, rather than 5-HT1AR, is the relevant 5-HTR for 5-HT-mediated inhibition of Ip in the present study. This still deserves further investigations.
Some studies indicate that overexpression of the 5-HT3R in mouse forebrain results in enhanced hippocampal-dependent learning and attention involved in fear conditioning , whereas most reports show that 5-HT3R antagonists can facilitate LTP and enhance the retention and consolidation of memory in hippocampal dependent tasks . Furthermore, the complete abolishment of 5-HT innervation in the hippocampus increases LTP in vivo -which would suggest that, on balance, 5-HT may exert a negative influence on LTP via 5-HT3R and then impair learning and memory [14, 15]. Clinically application of 5-HT3R antagonists have been found to improve memory in schizophrenic or Alzheimer demented patients [16, 17]. It is, however, not clear whether this effect is specific to LTP, or secondary to other changes.
Some studies reported that inhibition of Na+/K+ pump activity can induce LTD whereas depotentiate LTP , and then cause impairment of learning and memory and amnesia [19–21, 45–47]. Moreover, The initial stationary phase of the LTP was followed by a decrease in Na+/K+ pump activity of neurons and an augmentation of Na+/K+ pump activity in the glial cells . These studies supported that there may be some relationship between Na+/K+ pump and LTP. The present results show that 5-HT can suppress Ip in hippocampal CA1 pyramidal neurons via 5-HT3R, consistent with the previous study that 5-HT3R partly mediated the decrease of Na+/K+ pump activity induced by cocaine in neuronal-like cells , suggesting that inhibition of Na+/K+ pump activity may be involved in 5-HT3R-induced modulation of learning and memory. This provides new insights for the possible synaptic role of 5-HT via 5-HT3R in cognitive function and neuronal development through Na+/K+ pump, which may be a useful target to benefit these patients with cognitive disorder.
5-HT inhibits neuronal Na+/K+ pump activity via 5-HT3R in hippocampal CA1 pyramidal neurons, which may disclose novel mechanisms for the function of 5-HT in learning and memory.
Solutions and chemicals
Str, ondasetron and WAY-100635 were purchased from Sigma (St. Louis, MO, USA). 5-HT and tetrodotoxin (TTX) were purchased from Alexis (San Diego, CA, USA). m-CPBG and other chemicals were purchased from Alfa Aesar (Ward Hill, MA, USA). Str was dissolved in DMSO and further diluted 1: 1000 in artificial cerebrospinal fluid (ACSF) containing (mM): NaCl 119, KCl 5.4, MgCl2 1.3, NaH2PO4•2H2O 1, D-Glucose 11, NaHCO3 26.2, CaCl2 2.5. Control solutions of 1: 1000 DMSO had no effect on membrane current. TTX was dissolved in dilute acetic acid (PH 4.8-4.9). 5-HT, ondasetron, m-CPBG and WAY-100635 were dissolved in sterile water and stored as stock solutions. All stock solutions were stored as frozen aliquots at -20°C.
Brain hippocampal slice preparation and loading
Sprague-Dawley rats of 12-14 days were deeply anesthetized with sodium pentobarbital (45 mg kg-1, i.p.) and then rapidly decapitated. Our experiments were approved by Animal Care Committee of Hebei Medical University. Appropriate experimental procedures were taken to minimize pain or discomfort. The brain was quickly removed from the skull and transverse hippocampal slices (300 μm thick) were obtained by cutting with a vibroslice MA752 (Campden Instruments, Loughborough, UK) in ice-cold ACSF well-saturated with 95% O2 and 5% CO2 (PH 7.3-7.4). These slices were pre-incubated in oxygenated ACSF at room temperature (22-25°C) for 1 h.
Electrophysiological recordings of Na+, K+-pump currents
Results were expressed as the mean ± S.E.M., and n indicated the number of slices studied. Statistical comparisons were made using the Student's t-test for unpaired samples, and significant differences were defined as having a P-value less than 0.05.
This project was supported by the Natural Science Foundation of Hebei Province of China (No 301360).
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