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J M Bekkers

Publications and source records attributed to J M Bekkers.

At least 19 recordsLinked to original sources
PubMed2001

Modulation of excitability by alpha-dendrotoxin-sensitive potassium channels in neocortical pyramidal neurons.

Many neurons transduce synaptic inputs into action potentials (APs) according to rules that reflect their intrinsic membrane properties. Voltage-gated potassium channels, being numerous and diverse constituents of neuronal membrane, are important participants in neuronal excitability and thus in synaptic integration. Here we address the role of dendrotoxin-sensitive "D-type" potassium channels in the excitability of large pyramidal neurons in layer 5 of the rat neocortex. Low concentrations of 4-aminopyridine or alpha-dendrotoxin (alpha-DTX) dramatically increased excitability: the firing threshold for action potentials was hyperpolarized by 4-8 mV, and the firing frequency during a 1-sec-long 500 pA somatic current step was doubled. In nucleated outside-out patches pulled from the soma, alpha-DTX reversibly blocked a slowly inactivating potassium current that comprised approximately 6% of the total. This current first turned on at voltages just hyperpolarized to the threshold for spiking and activated steeply with depolarization. By assaying alpha-DTX-sensitive current in outside-out patches pulled from the axon and primary apical dendrite, it was found that this current was concentrated near the soma. We conclude that alpha-DTX-sensitive channels are present on large layer 5 pyramidal neurons at relatively low density, but their strategic location close to the site of action potential initiation in the axon may ensure that they have a disproportionate effect on neuronal excitability. Modulation of this class of channel would generate a powerful upregulation or downregulation of neuronal output after the integration of synaptic inputs.

4-Aminopyridine↗
PubMed2001

Inhibition of calcium channels by opioid- and adenosine-receptor agonists in neurons of the nucleus accumbens.

The pharmacological effects of opioid- and adenosine-receptor agonists on neural signalling were investigated by measuring drug actions on barium current flowing through calcium channels in acutely-dissociated neurons of the rat nucleus accumbens (NAc). Under whole-cell voltage clamp, opioids acted via mu, but not delta or kappa, receptors to partially inhibit barium current. Mean inhibition was 35+/-2% (+/-s.e.mean, n = 33) for methionine-enkephalin and 37+/-1% (n = 65) for the selective mu receptor agonist DAMGO, both measured at saturating agonist concentrations in neurons with diameter > or = 20 microm. EC(50) for DAMGO was 100 nM. Perfusion of naloxone reversed the current inhibition by DAMGO. Adenosine also partially inhibited barium current in these neurons. Mean inhibition was 28+/-2% (n = 29) for adenosine and 33+/-3% (n = 27) for the selective A1 receptor agonist N(6)CPA, both at saturating concentrations in neurons with diameter > or = 20 microm. EC(50) for N(6)CPA was 34 nM. Adenosine inhibition was reversed by perfusion of an A1 receptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine, while the selective A2A receptor agonist, CGS 21680, had no effect. Inhibition by opioids and adenosine was mutually occlusive, suggesting a converging pathway onto calcium channels. These actions involved a G-protein-coupled mechanism, as demonstrated by the partial relief of inhibition by strong depolarization and by the application of N-ethylmaleimide or GTP-gamma-S. Inhibition of barium current by opioids had their greatest effect in large neurons, that is, in presumed interneurons. In contrast, opioid inhibition in neurons with diameter < or = 15 microm was 11+/-2% (n = 26) for methionine-enkephalin and 11+/-4% (n = 17) for DAMGO, both measured at saturating agonist concentrations. Adenosine inhibition in neurons with diameter < or = 15 microm was 22+/-5% (n = 9). These results implicate the interneurons as a locus for the modulation of the excitability of projection neurons in the NAc during the processes of addiction and withdrawal.

Adenosine↗
PubMed2000

Properties of voltage-gated potassium currents in nucleated patches from large layer 5 cortical pyramidal neurons of the rat.

Voltage-gated potassium currents were studied in nucleated outside-out patches obtained from large layer 5 pyramidal neurons in acute slices of sensorimotor cortex from 13- to 15-day-old Wistar rats (22-25 C). Two main types of current were found, an A-current (IA) and a delayed rectifier current (IK), which were blocked by 4-aminopyridine (5 mM) and tetraethylammonium (30 mM), respectively. Recovery from inactivation was mono-exponential (for IA) or bi-exponential (for IK) and strongly voltage dependent. Both IA and IK could be almost fully inactivated by depolarising prepulses of sufficient duration. Steady-state inactivation curves were well fitted by the Boltzmann equation with half-maximal voltage (V ) and slope factor (k) values of -81.6 mV and -6.7 mV for IA, and -66.6 mV and -9.2 mV for IK. Peak activation curves were described by the Boltzmann equation with V and k values of -18.8 mV and 16.6 mV for IA, and -9.6 mV and 13.2 mV for IK. IA inactivated mono-exponentially during a depolarising test pulse, with a time constant ( approximately 7 ms) that was weakly dependent on membrane potential. IK inactivated bi-exponentially with time constants ( approximately 460 ms, approximately 4.2 s) that were also weakly voltage dependent. The time to peak of both IA and IK depended strongly on membrane potential. The kinetics of IA and IK were described by a Hodgkin-Huxley-style equation of the form mNh, where N was 3 for IA and 1 for IK. These results provide a basis for understanding the role of voltage-gated potassium currents in the firing properties of large layer 5 pyramidal neurons of the rat neocortex.

4-Aminopyridine↗
PubMed2000

Distribution and activation of voltage-gated potassium channels in cell-attached and outside-out patches from large layer 5 cortical pyramidal neurons of the rat.

Voltage-gated potassium channels were studied in cell-attached and outside-out patches from the soma and primary apical dendrite of large layer 5 pyramidal neurons in acute slices of rat sensorimotor cortex (22-25 degrees C). Ensemble averages revealed that some patches contained only fast, I(A)-like channels, other contained only I(K)-like channels that did not inactivate or inactivated slowly, and the remainder contained mixtures of both types. I(A) and I(K) channels had mean unitary conductances of 8.5 and 20.3 pS, respectively, and had distinctive patterns of gating. Peak activation curves for ensemble-averaged currents were described by the Boltzmann equation with half-maximal voltage [V(1/2)] and slope factor (k) values of -24.5 mV and 16.9 mV for I(A) and -7.6 mV and 10.1 mV for I(K) (patches < 250 microm from the soma) or -22.9 mV and 16.2 mV for I(A) (patches > 250 microm from the soma). The steady-state inactivation curve for I(A) gave V(1/2) and k values of -72.3 mV and -5.9 mV (< 250 microm from the soma) or -83.1 mV and -6.5 mV (> 250 microm from the soma). These values were similar to the corresponding data for I(A) and I(K) in nucleated patches from the same cell. The amount of I(A) and I(K) present in patches depended weakly on distance along the primary apical dendrite from the soma. The amplitude of I(A) increased, on the average, by 2.3 pA per 100 microm, while the amplitude of I(K) decreased by 0.4 pA per 100 microm. I(A) and I(K) channels in dendritic cell-attached patches were activated by the passage of a back-propagating action potential past the tip of the patch electrode. These results show directly that these potassium channels participate in action potential repolarisation, and thus contribute to the process of synaptic integration in these neurons.

Action Potentials↗
PubMed2000

Distribution of slow AHP channels on hippocampal CA1 pyramidal neurons.

This work was designed to localize the Ca(2+)-activated K(+) channels underlying the slow afterhyperpolarization (sAHP) in hippocampal CA1 pyramidal cells. Cell-attached patches on the proximal 100 microm of the apical dendrite contained K(+) channels, but not sAHP channels, activated by backpropagating action potentials. Amputation of the apical dendrite approximately 30 microm from the soma, while simultaneously recording the sAHP whole cell current at the soma, depressed the sAHP amplitude by only approximately 30% compared with control. Somatic cell-attached and nucleated patches did not contain sAHP current. Amputation of the axon >/=20 microm from the soma had little effect on the amplitude of the sAHP recorded in cortical pyramidal cells. By this process of elimination, it is suggested that sAHP channels may be concentrated in the basal dendrites of CA1 pyramids.

Action Potentials↗
PubMed1999

Quantal amplitude and quantal variance of strontium-induced asynchronous EPSCs in rat dentate granule neurons.

1. Excitatory postsynaptic currents (EPSCs) were recorded from granule cells of the dentate gyrus in acute slices of 17- to 21-day-old rats (22-25 C) using tissue cuts and minimal extracellular stimulation to selectively activate a small number of synaptic contacts. 2. Adding millimolar Sr2+ to the external solution produced asynchronous EPSCs (aEPSCs) lasting for several hundred milliseconds after the stimulus. Minimally stimulated aEPSCs resembled miniature EPSCs (mEPSCs) recorded in the same cell but differed from them in ways expected from the greater range of dendritic filtering experienced by mEPSCs. aEPSCs had the same stimulus threshold as the synchronous EPSCs (sEPSCs) that followed the stimulus with a brief latency. aEPSCs following stimulation of distal inputs had a slower mean rise time than those following stimulation of proximal inputs. These results suggest that aEPSCs arose from the same synapses that generated sEPSCs. 3. Proximally elicited aEPSCs had a mean amplitude of 6.7 +/- 2.2 pA (+/- s.d., n = 23 cells) at -70 mV and an amplitude coefficient of variation of 0. 46 +/- 0.08. 4. The amplitude distributions of sEPSCs never exhibited distinct peaks. 5. Monte Carlo modelling of the shapes of aEPSC amplitude distributions indicated that our data were best explained by an intrasite model of quantal variance. 6. It is concluded that Sr2+-evoked aEPSCs are uniquantal events arising at synaptic terminals that were recently invaded by an action potential, and so provide direct information about the quantal amplitude and quantal variance at those terminals. The large quantal variance obscures quantization of the amplitudes of evoked sEPSCs at this class of excitatory synapse.

Animals↗
PubMed1999

GABA(B), opioid and alpha2 receptor inhibition of calcium channels in acutely-dissociated locus coeruleus neurones.

1. The effects of GABA(B), opioid and alpha2 receptor activation on different subtypes of calcium channels in acutely-dissociated rat locus coeruleus (LC) neurones were investigated using whole-cell patch clamping. 2. Barium currents through calcium channels could be fractionated into four classes: L-type (nimodipine-sensitive), N-type (omega-conotoxin GVIA-sensitive), P/Q-type (omega-agatoxin IVA-sensitive) and R-type (remaining in the presence of all three blockers). The percentage of each was, respectively, 25+/-2, 34+/-1, 28+/-3 and 12+/-1% (mean+/-s.e.mean, n=4). 3. The GABA(B) receptor agonist, baclofen, and the opioid receptor agonist, enkephalin, partially inhibited the total barium current in a concentration-dependent manner with EC50 values of 2 and 0.3 microm , respectively. Maximal inhibition was 17+/-1% (n=38) for baclofen and 30+/-2% (n=20) for enkephalin. The alpha2-adrenoceptor agonist, UK14304 (10 microM), also inhibited barium current in these neurones (28+/-2%, n=11). The agonists did not shift the current-voltage relationship along the voltage axis. 4. Maximal baclofen inhibition of different calcium channel subtypes was 9+/-7% (L-type, n=4), 11+/-8% (N-type, n=4), 26+/-6% (P/Q-type, n=4), and 6+/-5% (R-type, n=5). The corresponding values for enkephalin inhibition were 5+/-9% (L-type), 30+/-11% (N-type), 37+/-9% (P/Q-type), and 17+/-8% (R-type). 5. In the presence of a saturating concentration of enkephalin, baclofen produced additional inhibition of the barium current. In contrast, in the presence of a saturating concentration of enkephalin, UK14304 produced no further inhibition of the barium current. 6. These results indicate that neuromodulation of calcium channels in LC neurones involves a complex pattern of overlapping and distinct second messenger pathways. Regulation of LC neuronal firing activity by the modulation of calcium channels may be important for LC-mediated behaviour such as alertness and vigilance.

Adrenergic alpha-2 Receptor Agonists↗
PubMed1998

N- and P/Q-type Ca2+ channels mediate transmitter release with a similar cooperativity at rat hippocampal autapses.

The relationship between extracellular Ca2+ concentration and EPSC amplitude was investigated at excitatory autapses on cultured hippocampal neurons. This relationship was steeply nonlinear, implicating the cooperative involvement of several Ca2+ ions in the release of each vesicle of transmitter. The cooperativity was estimated to be 3.1 using a power function fit and 3.3 using a Hill equation fit. However, simulations suggest that these values underestimate the true cooperativity. The role of different Ca2+ channel subtypes in shaping the Ca2+ dose-response relationship was studied using the selective Ca2+ channel blockers omega-agatoxin GIVA (omega-Aga), which blocks P/Q-type channels, and omega-conotoxin GVIA (omega-CTx), which blocks N-type channels. Both blockers broadened the dose-response relationship, and the Hill coefficient was reduced to 2.5 by omega-Aga and to 2.6 by omega-CTx. This broadening is consistent with a nonuniform distribution of Ca2+ channel subtypes across presynaptic terminals. The similar Hill coefficients in omega-Aga or omega-CTx suggest that there was no difference in the degree of cooperativity for transmitter release mediated via N- or P/Q-type Ca2+ channels. A model of the role of calcium in transmitter release is developed. It is based on a modified Dodge-Rahamimoff equation that includes a nonlinear relationship between extracellular and intracellular Ca2+ concentration, has a cooperativity of 4, and incorporates a nonuniform distribution of Ca2+ channel subtypes across presynaptic terminals. The model predictions are consistent with all of the results reported in this study.

6-Cyano-7-nitroquinoxaline-2,3-dione↗
PubMed1998

Neurophysiology: are autapses prodigal synapses?

Recent evidence suggests that 'autapses' - synapses made by a neuron with itself - are much more common in the brain than previously thought, leading to the possibility that they are more than just curiosities and may have a real physiological role.

Animals↗
PubMed1997

Nonuniform distribution of Ca2+ channel subtypes on presynaptic terminals of excitatory synapses in hippocampal cultures.

Several subtypes of Ca2+ channel support the release of glutamate at excitatory synapses. We investigated the pattern of colocalization of these subtypes on presynaptic terminals in hippocampal cultures. N-type (conotoxin GVIA-sensitive) or P/Q-type (agatoxin IVA-sensitive) Ca2+ channels were blocked selectively, and the reduction in transmitter release probability (Pr) was measured with MK-801. The antagonists completely blocked release at some terminals, reduced Pr at others, and failed to affect the remainder. In contrast, nonselective reduction of presynaptic Ca2+ influx by adding Cd2+ or lowering external Ca2+ reduced Pr uniformly at all terminals. We conclude from these results that the mixture of N-type and P/Q-type channels varies markedly between terminals on the same afferent. The distribution of Ca2+ channel subtypes was the same for high and low Pr terminals. Given that Ca2+ channel subtypes are affected differentially by neuromodulators, these findings lead to the possibility of terminal-specific modulation of synaptic function.

Animals↗
PubMed1997

Detection of spontaneous synaptic events with an optimally scaled template.

Spontaneous synaptic events can be difficult to detect when their amplitudes are close to the background noise level. Here we report a sensitive new technique for automatic detection of small asynchronous events. A waveform with the time course of a typical synaptic event (a template) is slid along the current or voltage trace and optimally scaled to fit the data at each position. A detection criterion is calculated based on the optimum scaling factor and the quality of the fit. An event is detected when this criterion crosses a threshold level. The algorithm automatically compensates for changes in recording noise. The sensitivity and selectivity of the method were tested using real and simulated data, and the influence of the template parameter settings was investigated. Its performance was comparable to that obtained by visual event detection, and it was more sensitive than previously described threshold detection techniques. Under typical recording conditions, all fast synaptic events with amplitudes of at least three times the noise standard deviation (3 sigma) could be detected, as could 75% of events with amplitudes of 2 sigma. The scaled template technique is implemented within a commercial data analysis application and can be applied to many standard electrophysiological data file formats.

Animals↗
PubMed1996

Apical dendritic location of slow afterhyperpolarization current in hippocampal pyramidal neurons: implications for the integration of long-term potentiation.

Trains of action potentials in hippocampal pyramidal neurons are followed by a prolonged afterhyperpolarization (AHP) lasting several seconds, which is attributable to the activation of a slow calcium-activated potassium current ((sI)AHP). Here we examine the location of (sI)AHP on CA1 pyramidal neurons by comparing it with two GABAergic inhibitory postsynaptic currents (IPSCs) with known somatic and dendritic locations. Whole-cell patch-clamp recordings were made for CA1 pyramidal neurons in acute hippocampal slices. Stepping the membrane potential at the peak of (sI)AHP produced a relaxation ("switchoff") of the AHP current with a time constant of 7.4 +/- 0.4 msec (mean +/- SEM). The switchoff time constants for somatic and dendritic GABAA IPSCs were 3.5 +/- 0.5 msec and 8.8 +/- 0.3 msec, respectively. This data, together with cable modeling, indicates that active (sI)AHP channels are distributed over the proximal dendrites within approximately 200 micrometers of the soma. Excitatory postsynaptic potentials (EPSPs) evoked in stratum (s.) radiatum had their amplitudes shunted more by the AHP than did EPSPs evoked in s. oriens, suggesting that active AHP channels are restricted to the apical dendritic tree. Blockade of the AHP during a tetanus, which in control conditions elicited a decremental short-term potentiation (STP), converted STP to long-term potentiation (LTP). Thus, activation of the AHP increases the threshold for induction of LTP. These results suggest that in addition to its established role in spike frequency adaptation, the AHP works as an adjustable gain control, variably hyperpolarizing and shunting synaptic potentials arising in the apical dendrites.

Action Potentials↗
PubMed1996

Differential effects of histamine on the N-methyl-D-aspartate channel in hippocampal slices and cultures.

The effect of histamine on N-methyl-D-aspartate currents was investigated in pyramidal neurons in the CA1 region of acute hippocampal slices from juvenile rats. The objective was to compare histamine effects in the slice with those previously reported in acutely dissociated and cultured hippocampal neurons. Micromolar concentrations of histamine had no effect on N-methyl-D-aspartate mediated excitatory postsynaptic currents in the slice, in contrast to the large enhancement seen in culture under identical conditions. However, millimolar concentrations of histamine blocked these currents both in the slice and in culture. Possible reasons for the lack of enhancement in the slice were explored as follows. (1) Histamine could not penetrate the slice or was already present at high concentrations inside the slice. This was tested by recording N-methyl-D-aspartate currents elicited in outside-out patches pulled from the somas of CA1 slice neurons. Histamine still had no effect in patches, whereas the corresponding experiment for cultured neurons showed robust enhancement. (2) Slices release an endogenous ligand that binds with high affinity to the histamine site on the N-methyl-D-aspartate receptor, blocking its activation. This was tested by superfusing cultures with supernatant from homogenized slice tissue. Histamine enhancement was maintained in these cultures. (3) CA1 slices and cultures express different N-methyl-D-aspartate receptor subtypes. The reverse transcription-polymerase chain reaction technique was used to examine the expression of messenger RNA encoding N-methyl-D-aspartate receptor subunits in the two systems. No difference was found in the whole-tissue expression of messenger RNA for the NR2A, 2B or 2C subunits or for the eight known splice variants of the NR1 subunit. It is hypothesized that the differential enhancing effect of histamine in slices and culture involves posttranslational modifications or other factors that modulate the N-methyl-D-aspartate receptor/ion channel according to its environment.

Animals↗
PubMed1996

Cable properties of cultured hippocampal neurons determined from sucrose-evoked miniature EPSCs.

1. The passive cable properties of rat hippocampal neurons in dissociated culture were studied using focal application of hypertonic solution to locally elicit miniature excitatory postsynaptic currents (mEPSCs) on the soma and dendrites. Neurons were filled with Lucifer yellow and portions of their dendritic trees were measured. 2. The average mEPSC measured at the soma appeared smaller and slower as the site of sucrose application was made more distal. Normalizing to a 1-micron diam dendrite, the mean mEPSC peak amplitude and charge was reduced e-fold in 170 and 1,000 microns, respectively, and the mean mEPSC decay time constant was increased e-fold in 150 microns. However, for any particular sucrose site, individual mEPSCs varied widely in their amplitudes and time courses. Plots of individual peak amplitudes versus half-width or rise time showed much overlap for mEPSCs originating from sites as much as 100 microns apart. This suggests that use of such plots to estimate the electrotonic location of synaptic currents is highly prone to error. 3. Averaged mEPSCs recorded when applying sucrose at the soma were poorly fitted by an alpha function but were well-described by an equation of the form mxh, where m incorporates a rise-time constant tau 1 and h a decay time constant tau 2. Averaged fits to mean mEPSCs elicited at the somas of five cells gave (mean +/- SE): peak conductance = 832 +/- 126 pS, tau 1 = 0.29 +/- 0.06 ms, tau 2 = 3.03 +/- 0.24 ms, x = 4.7 +/- 0.7. 4. For three cells, the entire dendritic branch to which sucrose was applied was measured and used to construct a passive cable model. The specific membrane resistance (Rm) and intracellular resistivity (Ri) were varied systematically in the model (assuming membrane capacitance Cm = 1 microF/cm2) to search for the best agreement between the mean mEPSCs and the model. Optimal Rm was found to lie in the range 20-30 k omega cm2, Ri in the range 100-200 omega cm. 5. These results confirm those obtained by other methods and emphasize the considerable cable filtering of fast electrical events in cultured hippocampal neurons.

Animals↗
PubMed1995

Quantal analysis of EPSCs recorded from small numbers of synapses in hippocampal cultures.

1. We have studied the statistical properties of excitatory post-synaptic currents (EPSCs) measured at small numbers of synaptic contacts between pairs of hippocampal neurons maintained in dissociated cell culture. Synaptic transmission at few synapses was enabled by microperfusion of a small region of the postsynaptic cell with Ca-containing solution, while blocking transmission at all other synaptic boutons by bathing them in low-Ca solution. Frequency histograms of the amplitudes of EPSCs recorded in this way showed no clear quantization. Numbers of active synapses, estimated immunohistochemically with the use of light microscopy, ranged from 4 to 14 in different experiments. 2. Miniature EPSCs (mEPSCs), originating in the same small population of synapses as produced the evoked EPSCs, were elicited by microperfusion of bath solution made hypertonic by the addition of sucrose. These "sucrose-evoked" mEPSCs appeared to be identical to "spontaneous" mEPSCs in every respect except control over their frequency and site of origin. Sucrose-evoked mEPSCs originating in few synapses still exhibited a broad amplitude distribution. Thus, if mEPSCs constitute the postsynaptic response to a single quantum of neurotransmitter (the "quantal amplitude"), their broad amplitude distribution would tend to obliterate evidence of quantization in evoked EPSC amplitudes, even if evoked release was, indeed, quantal. 3. This idea, which is a corollary of the Katz model of quantal transmission, was tested quantitatively by assuming 1) neurotransmitter release obeys uniform binomial statistics, and 2) the quantal amplitude has a distribution given by the observed distribution of sucrose-evoked mEPSCs. The expected distribution, calculated on the basis of these two assumptions, was fitted to the observed distribution of evoked EPSC amplitudes by varying two free parameters, the binomial parameters N and p. In five cells out of six that were fully analyzed, the Poisson limit of the binomial model (N large, p small) provided a very good fit to the data. This and other evidence suggests that the release probability at a single presynaptic terminal is low. In two out of the six cells, the binomial model, with N constrained to the histochemically determined bouton count, yielded acceptable fits; for the remaining cells the constrained binomial model could be rejected. 4. It is concluded that the Katz model of quantized release of neurotransmitter gives an adequate description of excitatory synaptic transmission in hippocampal cultures, when one assumes the broad distribution of mEPSC amplitudes reflects the distribution of the postsynaptic effect of a single quantum of transmitter.

Animals↗
PubMed1994

Quantal analysis of synaptic transmission in the central nervous system.

In the past year, important advances have been made in the understanding of quantal neurotransmission at central synapses. These include new statistical tests for the significance of quantal peaks in synaptic amplitude histograms, greater understanding of possible sources of quantal variance, and new attempts to undertake a rigorous quantal analysis of neurotransmission.

Animals↗
PubMed1993

Enhancement by histamine of NMDA-mediated synaptic transmission in the hippocampus.

Histamine is a neuromodulator in the brain, and the hippocampus is one of the regions of the brain that is innervated by histaminergic neurons. When applied to cultured hippocampal neurons, histamine selectively increased by up to tenfold the amplitude of the component of synaptic transmission that was mediated by N-methyl-D-aspartate (NMDA) receptors. Spontaneous miniature synaptic currents and the current elicited by applied NMDA also were enhanced, indicating that the histamine effect was expressed primarily postsynaptically. These results suggest that histamine may modulate processes involving NMDA receptors, such as the induction of long-term potentiation.

Animals↗