Guinea Pig Hippocampal CA1 Neurons Research Articles

Role of postsynaptic inositol 1, 4, 5-trisphosphate receptors in depotentiation in guinea pig hippocampal CA1 neurons

The long-term potentiation (LTP) in the field excitatory postsynaptic potential (EPSP) induced at hippocampal CA1 pyramidal neuron synapses by delivery of high frequency stimulation (HFS), a tetanus of 100 pulses at 100Hz, is decreased (depotentiation) by a train of low frequency stimulation (LFS) of 1000 pulses at 2Hz applied 30min later. Inositol 1, 4, 5-trisphosphate receptors (IP3Rs) activated both during the HFS and after the LFS are involved in this depotentiation, the former triggering, and the latter modifying, LTP induction (decreasing the amplitude of the LTP established by the priming HFS). Furthermore, the decrease in the LTP at CA1 synapses requires activation of IP3Rs during LFS and activation of calcineurin after LFS. These results suggest that, at hippocampal CA1 neuron synapses, HFS-induced IP3R activation, which is modulated by the subsequent LFS, results in postsynaptic protein dephosphorylation after the LFS, leading to a decrease in the field EPSP and in the HFS-induced LTP.

Involvement of IP3 receptors in LTP and LTD induction in guinea pig hippocampal CA1 neurons

The role of inositol 1, 4, 5-trisphosphate receptors (IP3Rs) in long-term potentiation (LTP) and long-term depression (LTD) was studied in CA1 neurons in guinea pig hippocampal slices. In standard solution, short tetanic stimulation consisting of 15 pulses at 100 Hz induced LTP, while three short trains of low-frequency stimulation (LFS; 200 pulses at 1 Hz) at 18-min intervals or one long train of LFS (1000 pulses at 1 Hz) induced stable LTD in both the slope of the field EPSP (S-EPSP) and the amplitude of the population spike (A-PS). Bath application of 2-aminoethoxydiphenyl borate (2-APB), an IP3R antagonist, or of alpha-methyl-4-carboxyphenylglycine (MCPG), a wide-spectrum metabotropic glutamate receptor antagonist, during weak tetanic stimulation significantly increased the magnitude of the LTP in both the S-EPSP and A-PS. Three short trains of LFS or one long train of LFS delivered in the presence of 2-APB or MCPG did not induce LTD, but elicited LTP. Based on these results, we conclude that, in hippocampal CA1 neurons, IP3Rs play an important role in synaptic plasticity by attenuating LTP and facilitating LTD.

Long‐term potentiation and long‐term depression induced by local application of ATP to hippocampal CA1 neurons of the Guinea pig

The present study has investigated the role of ATP in the induction of synaptic plasticity, using local application of ATP by picopump administration into the stratum radiatum of guinea pig hippocampal region CA1. Excitatory postsynaptic currents (EPSCs) evoked by stimulation of Schaffer collateral/commissural afferents synapsing on CA1 pyramidal cells of hippocampal slices were monitored in voltage-clamp mode, using whole-cell recording. Brief local application of ATP (1 mM) induced an inward current, usually consisting of early- and late-phase components. Because the late-phase component of an ATP-induced current was largely inhibited by Ca2+-free solution, this component is supposed to depend on extracellular Ca2+. After local application of ATP, long-term synaptic modification of EPSCs was induced: LTP was detected in neurons exhibiting a small late Ca2+ current, while LTD was obtained from recordings showing a large late Ca2+ current in response to ATP application. There was a statistically significant correlation between the magnitude of long-term plastic changes and the size of Ca2+ currents in response to ATP application. Furthermore, there was significant difference between the average size of the Ca2+ current in the LTP group and the size in the LTD group. These results suggest that a small Ca2+ influx in response to ATP application induces LTP, whereas a large one induces LTD in guinea pig hippocampal CA1 neurons.

Tetanus-induced potentiation of inhibitory postsynaptic potentials in hippocampal CA1 neurons.

In the present study, tetanus-induced potentiation of inhibitory postsynaptic potentials (IPSPs), previously described by our laboratory, was further investigated in guinea pig hippocampal CA1 neurons. Tetanic stimulation of the stratum radiatum induced a long-term potentiation of the excitatory postsynaptic potential and a potentiation of the gamma-aminobutyric acid A (GABAA) receptor mediated fast IPSP without enhancing the GABAB receptor-mediated slow IPSP. During the potentiation, IPSPs evoked by stimulation of the alveus were unaffected. When slices were superfused with DL-2-amino-5-phosphonovaleric acid (an N-methyl-D-aspartate, NMDA, antagonist) and 6-cyano-7-nitroquinoxaline-2,3-dione (a non-NMDA glutamate antagonist), the potentiation of the monosynaptic fast IPSP could still be induced and maintained, suggesting that polysynaptic influences were unnecessary for this process. Finally, since the potentiation was observed in CA1 neurons in which BAPTA or K-252b was injected, this form of plasticity does not appear to be dependent on a rise in intracellular [Ca2+] or protein kinase C (PKC) activity. These results indicate that tetanic stimulation of the stratum radiatum induces a potentiation of GABAergic fast IPSPs in CA1 neurons. The potentiation may be localized to the GABAergic synapse on CA1 neurons.

The mechanism of ATP-induced long-term potentiation involves extracellular phosphorylation of membrane proteins in guinea-pig hippocampal CA1 neurons

The mechanism of ATP-induced long-term potentiation was studied pharmacologically using guinea-pig hippocampal slices. Application of 1–10 μM ATP for 10 min transiently depressed and then slowly augmented the synaptic transmission in CA1 neurons leading to long-term potentiation (LTP). This ATP-induced LTP was blocked by the addition of K-252b, an ecto-protein kinase inhibitor, but was enhanced by the addition of RK682, an ecto-phosphatase inhibitor, both of which do not permeate the cell membrane. These results suggest that ATP applied to the perfusate provides enough substrate for ecto-protein kinase to induce LTP through phosphorylation of extracellular domains of membrane proteins in CA1 neurons.

Voltage-gated Ca 2+ channel blockers, ω-AgalVA and Ni 2+, suppress the induction of θ-burst induced long-term potentiation in guinea-pig hippocampal CA1 neurons

It is widely believed that a rise in post-synaptic calcium concentration ([Ca 2+];) is a necessary step in the induction of long-term potentiation (LTP) (Bliss and Collingridge, Nature, 361 (1993) 31–39). In this experiment, we examine the involvement of voltagegated Ca 2+ channels (VGCC) in the induction of AP5-sensitive LTP induced by θ-burst stimulation in guinea-pig hippocampal CA1 neurons. The VGCC blockers, Ni 2+, (25 μM, T -channel blocker) or ω-AgaIVA (60 nM, P-channel blocker), which have no effect on synaptic transmission, suppress 60% or 78% of the Θ-burst induced UP, respectively. This implies that Ca 2+ entry through VGCC is an important step in this form of LTP.

Cortisol inhibition of calcium currents in guinea pig hippocampal CA1 neurons via G-protein-coupled activation of protein kinase C

The inhibition of voltage-activated Ca2+ channel currents by cortisol (hydrocortisone), the principal glucocorticoid in man and guinea pig, was examined in freshly dissociated pyramidal neurons from the adult guinea pig hippocampal CA1 region using whole-cell voltage-clamp recordings. Steady-state inhibition by cortisol of the peak Ca2+ channel current evoked by depolarization from -80 to -10 mV increased in a concentration-dependent fashion, with a maximal inhibition of 63 +/- 4% of the total current at 100 microM. Cortisone had a maximal 17 +/- 2% inhibition at 10 microM. Corticosterone and the metabolite allotetrahydrodeoxycorticosterone exhibited a plateau of inhibition of around 15% and 25%, respectively, between 10 pM and 100 nM; both compounds continued to inhibit at concentrations > 10(-7) M. Analysis of tail currents at -80 mV showed that cortisol and corticosterone had no effect on the voltage-dependent activation or deactivation of the Ca2+ channel current. However, cortisol slowed the activation of the current. Cortisol inhibited both the N-type or omega-conotoxin (CgTX)-sensitive, and the L-type or nifedipine (NIF)-sensitive Ca2+ channel current but had no effect on the CgTX/NIF-insensitive Ca2+ channel current. In neurons isolated from pertussis toxin (PTX)-treated animals, the cortisol inhibition was significantly diminished. Intracellular dialysis with GDP-beta-S (500 microM) or with the specific inhibitors of protein kinase C (PKC), the pseudosubstrate PKC inhibitor (PKCI 19-31) (2 microM) and bisindolylmaleimide (BIS) (1 microM) significantly diminished the cortisol inhibition of the Ca2+ channel current. The specific inhibitor of cAMP-dependent protein kinase (PKA) inhibitor, Rp-cAMPS (100 microM) had no effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin-1β inhibits Ca 2+ channel currents in hippocampal neurons through protein kinase C

Interleukin-1β depresses the voltage-gated Ca 2+ channel currents in acutely dissociated guinea-pig hippocampal CA1 neurons. This depression is observed with pathophysiological concentrations found in the cerebrospinal fluid (≥ 1.0 pg interleukin-1 β/10 μl). Interleukin-1 receptor antagonist (in concentrations 25-fold higher than interleukin-1β) completely blocked the interleukin-1β-induced depression of the Ca 2+ channel current. This suggests that interleukin-1β action is through a specific interaction with an interleukin-1 membrane receptor site. The application of other cytokines and growth factors (interleukin-6, epidermal growth factor, and basic fibroblast growth factor), or bacterial lipopolysaccharide (endotoxin) had no effect, indicating specificity of action of interleukin-1β. The depression of the Ca 2+ channel current by interleukin-1β was prevented by the extracellular application of pertussis toxin, and by the intracellular application of GDP[βS], H-7, staurosporine or bisindolylmaleimide. Application of phorbol 12-myristate 13-acetate also depressed the Ca 2+ channel current, but this phorbol ester-induced depression was not additive to that induced by interleukin-1β. These results suggest mediation of interleukin-1β action through a pertussis toxin-sensitive G-protein coupled interleukin-1 receptor associated with the activation of protein kinase C. The depression of the Ca 2+ channel current by interleukin-1β may be involved in the regulation of neuronal excitability during pathological conditions and in the induction and/or progression of neurodegenerative processes.

Calcium current block by (-)-pentobarbital, phenobarbital, and CHEB but not (+)-pentobarbital in acutely isolated hippocampal CA1 neurons: comparison with effects on GABA-activated Cl- current

Block of a voltage-activated Ca2+ channel current by phenobarbital (PHB), 5-(2-cyclohexylideneethyl)-5-ethyl barbituric acid (CHEB), and the optical R(-)- and S(+)-enantiomers of pentobarbital (PB) was examined in freshly dissociated adult guinea pig hippocampal CA1 neurons; the effects of the barbiturates on GABA-activated Cl- current were also characterized in the same preparation. (-)-PB, PHB, and CHEB produced a reversible, concentration-dependent block of the peak Ca2+ channel current (3 mM Ba2+ as the charge carrier) evoked by depolarization from -80 to -10 mV (IC50 values, 3.5, 72, and 118 microM, respectively). In contrast, (+)-PB was nearly inactive at concentrations up to 1 mM. The inhibitory action of PHB was decreased at acid pH, indicating that the dissociated (anionic) form of the molecule is the active species. Block by (-)-PB was voltage dependent with the fractional block increasing at positive membrane potentials; calculations according to the method of Woodhull indicated that the (-)-PB blocking site senses approximately 40% of the transmembrane electric field. The time course and voltage dependence of activation of the Ca2+ channel current were unaffected by (-)-PB, PHB, and CHEB. The rate of inactivation was enhanced by (-)-PB and CHEB, with the major effect being acceleration of the slow phase of the biexponential decay of the current. GABA-activated Cl- current was potently enhanced by (-)-PB and PHB (EC50 values, 3.4 and 12 microM), whereas (+)-PB was only weakly active. At concentrations of (-)-PB > 100 microM and PHB > 200-300 microM, Cl- current responses were activated even in the absence of GABA. These results demonstrate that in CA1 hippocampal neurons, PB causes a stereoselective block of a voltage-activated Ca2+ current; PHB is also effective, but at higher concentrations. For (-)-PB, the effect on Ca2+ channel current occurred at similar concentrations as potentiation of GABA responses. In contrast, PHB was more potent as a GABA enhancer than as blocker of Ca2+ current, but the maximal potentiation of GABA responses was 40% of that obtained with (-)-PB. Consequently, the anticonvulsant action of PHB at clinically relevant concentrations may relate to modest enhancement of GABA responses and partial blockade of Ca2+ current, whereas the sedative effects that occur at higher concentrations could reflect stronger Ca2+ current blockade. The powerful sedative-hypnotic action of (-)-PB may reflect greater maximal enhancement of GABA responses in conjunction with strong inhibition of Ca2+ current.(ABSTRACT TRUNCATED AT 400 WORDS)

Interleukin-2 modulates calcium currents in dissociated hippocampal CA1 neurons

Interleukin-2 and its receptors are present in hippocampal structures, particularly the pyramidal cell layer. In the present study, we examined the interaction between recombinant human interleukin-2 (rhIL-2) and the voltage-dependent calcium (Ca2+) current using the whole-cell patch-clamp technique. RhIL-2 depressed a fraction of the inward Ca2+ current in acutely dissociated guinea-pig hippocampal CA1 neurons. This depression is rapid, dose-dependent with 0.001-1.0 ng rhIL-2/10 microliters (6.8 x 10(-12) to 6.8 x 10(-9) M) and reversible in > 80% of the neurons. Heat-inactivated rhIL-2 had no effect. The application of anti-rhIL-2 monoclonal antibody inhibited the rhIL-2-induced depression of the Ca2+ current. The depression of the Ca2+ current by IL-2 may play a role in the induction and/or progression of neuropathological processes of immunological origin, and in the induction of neurological manifestations during IL-2 immunotherapy.

Induction of hippocampal long-term potentiation by α-tocopherol

Long-term potentiation (LTP) of synaptic transmission in the hippocampus is thought to be one of the cellular mechanisms underlying learning and memory. Recent evidence in literature suggests the involvement of free radicals in impeding LTP maintenance. In the present study, the effects of α-tocopherol, a major lipid-soluble antioxidant which could prevent lipid peroxidation, were examined on the excitatory post-synaptic potentials (EPSPs) of CA1 neurons in guinea pig hippocampal slices. α-Tocopherol phosphate disodium salt (0.2 mM applied for 5 min) induced a slowly developing long-lasting increase of the EPSP, without significantly changing the membrane potential, the input resistance and the ability to generate action potentials. No significant changes in the fast and the slow inhibitory post-synaptic potentials (IPSPs) were observed during the α-tocopherol-induced LTP of the EPSP. 2-Amino-5-phosponovalerate (APV) did not block the induction of this LTP. l-Ascorbic acid (Na salt, 3–10 mM), a water-soluble antioxidant, failed to produce any significant enhancement in the EPSP. These results indicate that α-tocopherol can induce LTP of the EPSP in guinea pig hippocampal CA1 neurons. The activation of N-methyl- d-aspartate (NMDA) receptors does not appear to be necessary for this action of α-tocopherol. Whether the LTP-inducing action of this agent is related to its antioxidant property is unclear.

Interleukin-1β depresses calcium currents in CA1 hippocampal neurons at pathophysiological concentrations

Interleukin-1 is present in the central nervous system (CNS) during acute and chronic pathological processes. In the present study, we examined the interaction between recombinant human interleukin-1β (rhIL-1β) and the voltage-dependent calcium (Ca 2+) current using the whole-cell patch clamp technique. RhIL-1β depressed the voltage-gated Ca 2+ current in acutely dissociated guinea pig hippocampal CA1 neurons. This depression is rapid and is observed at pathophysiological concentrations (⩾ 1.97 pg/10 μl)- Concomitant application of rhIL-1β and rhIL-1 receptor antagonist had no effect indicating neuroactive specificity of rhIL-1β. The depression of the inward Ca 2+ current by IL-1β may play a role in: 1. 1) the regulation of neuronal excitability; 2. 2) the induction of neurological manifestations during disease; and 3. 3) in the induction and/or progression of neurodegenerative processes.

The neurosteroids pregnenolone and pregnenolone-sulfate but not progesterone, block Ca 2+ currents in acutely isolated hippocampal CA1 neurons

The neurosteroids pregnenolone (PE) and pregnenolone-sulfate (PS) have been shown to interact with the GABA A receptor in the central nervous system. In contrast, nothing is known of any possible modulation of voltage-gated calcium channels (VGCC). We have examined the interaction of PE, PS and progesterone on VGCC in acutely isolated adult guinea-pig hippocampal CA1 neurons using the whole-cell patch clamp technique. PE and PS depressed the calcium current at low micromolar concentrations (0.001 – 100 μM). The time to peak of the calcium current was slowed by PE and PS. The blocking action of PE and PS occurs in the presence of 10 μM picrotoxin. In contrast, progesterone had no effect on the Ca 2+ current, indicating specificity for PE and PS. These results demonstrate a direct and novel membrane site of action for PE and PS, suggesting a possible role influencing brain excitability.

Phencyclidine actions measured intracellularly in hippocampal CA1 neurons

The electrophysiological effects of phencyclidine (PCP) were measured intracellularly in guinea pig hippocampal CA1 neurons in vitro. At all doses tested (0.2 microM - 10 mM), PCP increased the width of action potentials (APs). Doses of 10 microM and higher were associated with decreased action potential amplitude. PCP decreased inhibitory postsynaptic potentials and excitatory postsynaptic potentials but did not alter responses to focally applied GABA. At the lowest dose (0.2 microM), PCP decreased the input resistance (Rin), while at all other doses Rin was increased. PCP decreased post-spike train afterhyperpolarizations at low and medium doses. PCP effects persisted in low calcium medium and also in medium containing 10(-6) M tetrodotoxin. It is concluded that in these central neurons, PCP primarily blocks potassium conductances at all doses and, at anesthetic doses, depresses sodium-dependent spikes.

Calcium-dependent potentials with different sensitivities to calcium agonists and antagonists in guinea-pig hippocampal neurons

Effects of organic Ca channel blockers, Ca channel activators and ω-conotoxin on guinea-pig hippocampal CA1 neurons in in vitro preparations were studied with intracellular recording methods. Most of the Ca channel blockers, such as prenylamine, D 600, flunarizine, nifedipine, cinnarizine and nicardipine (0.2–4 μM), raised the threshold for Na-dependent spike generation and decreased the amplitude of the spike afterhyperpolarization. Verapamil (5 μM) and diltiazem (5 μM) did not significantly alter the threshold and amplitude of the Na spike. Action potentials elicited in the presence of either tetrodotoxin (0.5 μM) and tetraethylammonium (20 mM) or tetrodotoxin (0.5 μM) and Ba (1.25 mM) consisted of an initial spike component followed by a long depolarization. Both responses were abolished by addition of Co (2 mM) or Cd (0.25–0.5 mM), or by superfusion with a low Ca (0.25 mM)-high Mg(15 mM) medium, indicating that the potentials resulted from Ca entry. The Ca-dependent slow depolarization was preferentially blocked by most of the organic Ca channel blockers at approximately one-third the concentrations (0.1–2 μM) which were required to shorten the Ca spike. When the cell in a solution containing tetrodotoxin (0.5 μM), Co (2 mM) and 4-aminopyridine (2 mM) was hyperpolarized and then depolarized by passing current pulses across the membrane, a transient depolarizing hump occurred on the decay phase of the electrotonic potential. This transient depolarization was abolished by Co (2 mM), Ni (2 mM) or most of the organic Ca channel blockers (0.2–5 μM). Diltiazem (5 μM) did not significantly change these Ca-dependent potentials. The evoked excitatory postsynaptic potential was very resistant to the Ca channel blockers. Approximately 2–10 times higher concentrations (0.5–3 μM) were necessary to decrease the excitatory postsynaptic potential amplitude than to shorten the Ca spike. On the other hand, the minimal concentrations and order of potencies of the Ca channel blockers for depressing the evoked inhibitory postsynaptic potential and for elevating the threshold for Na spike generation were almost the same. Dihydropyridine Ca channel activators, such as Bay K 8644, CGP 28 392 and YC 170 at low concentrations (0.1–1 μM), decreased the Ca spike, the Ca-dependent slow depolarization and the evoked synaptic potentials, while the substances augmented the Ca-dependent transient depolarization. On the other hand, ω-conotoxin (5 μM) reversibly depressed the Ca spike and slow depolarization to the same degree, without affecting the transient depolarization and the evoked excitatory or inhibitory postsynaptic potentials. From these results, subtypes of Ca channels involved in generating the Ca-dependent responses and synaptic potentials have been discussed.

The sodium current underlying action potentials in guinea pig hippocampal CA1 neurons.

Neurons were acutely dissociated from the CA1 region of hippocampal slices from guinea pigs. Whole-cell recording techniques were used to record and control membrane potential. When the electrode contained KF, the average resting potential was about -40 mV and action potentials in cells at -80 mV (current-clamped) had an amplitude greater than 100 mV. Cells were voltage-clamped at 22-24 degrees C with electrodes containing CsF. Inward currents generated with depolarizing voltage pulses reversed close to the sodium equilibrium potential and could be completely blocked with tetrodotoxin (1 microM). The amplitude of these sodium currents was maximal at about -20 mV and the amplitude of the tail currents was linear with potential, which indicates that the channels were ohmic. The sodium conductance increased with depolarization in a range from -60 to 0 mV with an average half-maximum at about -40 mV. The decay of the currents was not exponential at potentials more positive than -20 mV. The time to peak and half-decay time of the currents varied with potential and temperature. Half of the channels were inactivated at a potential of -75 mV and inactivation was essentially complete at -40 to -30 mV. Recovery from inactivation was not exponential and the rate varied with potential. At lower temperatures, the amplitude of sodium currents decreased, their time course became longer, and half-maximal inactivation shifted to more negative potentials. In a small fraction of cells studied, sodium currents were much more rapid but the voltage dependence of activation and inactivation was very similar.

The influence of dopamine of epileptiform burst activity in hippocampal pyramidal neurons

Dopamine (DA) application to guinea pig hippocampal CA1 neurons in vitro causes hyperpolarization of the resting potential, increase in conductance, and increase in amplitude and duration of the afterhyperpolarization (AHP). Since these changes could influence repetitive firing, we performed experiments to determine whether DA-induced effects would suppress epileptogenesis in the hippocampus. Epileptiform bursts were induced by adding penicillin (3.4 mM) to the perfusion medium. Focal application of DA (40–160 μm) onto CA1 cells (n = 15) produced a hyperpolarization averaging 4.5 mV beginning in 5–20 s and lasting up to 3 min. DA also caused an increase in the amplitude and duration of slow AHPs. The frequency of spontaneous epileptiform events however was not affected. CA3 neurons (n = 6) responded to DA application with an initial 1–3 mV depolarization beginning within 5–30 s and lasting 1–2 min. In 3 cases a small hyperpolarization lasting several minutes subsequently developed. AHP duration increased 70% and amplitude increased 35% (n = 4). Along with these membrane changes the frequency of epileptiform bursting in CA3 cells slowed for 1–3 min. We added DA (10–80 μM) to the perfusion medium to see whether a significant decrease in epileptiform burst frequency might occur in the follower CA1 region if greater numbers of pacemaker CA2 and CA3 cells were exposed to DA. Spontaneous CA1 bursting was reversibly slowed, the interburst interval became variable and increased from a mean of 4 to a mean of 5–7 s (n = 6). These results suggest that DA may play a role in decreasing the incidence or frequency of epileptogenic discharges in vivo.