Large amplitude miniature excitatory postsynaptic currents in hippocampal CA3 pyramidal neurons are of mossy fiber origin

Neonatal (P0) gamma-irradiation was used to lesion selectively the mossy fiber (MF) synaptic input to CA3 pyramidal cells. This lesion caused a > 85% reduction in the MF input as determined by quantitative assessment of the number of dynorphin immunoreactive MF boutons. The gamma-irradiation lesion caused a reduction in the mean number of miniature excitatory postsynaptic currents (mEPSCs) recorded from CA3 pyramidal cells (2,292 vs. 1,429/3-min period; n = 10). The lesion also caused a reduction in the mean mEPSC peak amplitude from 19.1 +/- 0.45 to 14.6 +/- 0.49 pA (mean +/- SE; peak conductance 238.8 +/- 5.6 to 182.0 +/- 6.1 pS). Similarly, there was a reduction in the mean 10-85% rise time from 1.72 +/- 0.02 ms to 1.42 +/- 0.04 ms. The effects of the gamma-irradiation on both mEPSC amplitude and 10-85% rise time were significant at P < 0.002 and P < 0.005 (2-tailed Kolmogorov-Smirnov test). Based on the selectively of the gamma-irradiation, MF and non-MF mEPSC amplitude and 10-85% rise-time distributions were calculated. Both the amplitude and 10-85% rise-time distributions showed extensive overlap between the MF and non-MF mediated mEPSCs. The MF mEPSC distributions had a mean peak amplitude of 24.6 pA (307.5 pS) and a mean 10-85% rise time of 2.16 ms. THe non-MF mEPSC distributions had a mean peak amplitude of 12.2 pA (152.5 pS) and 10-85% rise time of 1.26 ms. The modes of the amplitude distributions were the same at 5 pA (62 pS). The MF and non-MF mEPSC amplitude and 10-85% rise-time distributions were significantly different at P < 0.001 (1-tailed, large sample Kolmogorov-Smirnov test). The data demonstrate that the removal of the MF synaptic input to CA3 pyramidal cells leads to the absence of the large amplitude mEPSCs that are present in control recordings.     

Activity-dependent response depression in rat hippocampal CA1 pyramidal neurons in vitro

1. A gradual and prolonged decrease of the response, termed here "depression," evoked by repeated activation with transmembrane current stimuli was analyzed in rat CA1 hippocampal pyramidal cells under single-electrode current clamp by the use of the in vitro slice technique. 2. Depression was induced by 2-s duration 0.3- to 0.7-nA current pulses presented as a sequence of 12 stimuli at 3- to 60-s intervals. Sinusoidal currents (0.5-1.0 nA) at 5-Hz or 200-ms pulses repeated at 0.3-0.5/s, which may be more natural stimulations, also induced depression. 3. Depression outlasted stimulation up to 170 s in all cells tested. The initial high rate spike burst changed little (< 20%), whereas the lower rate adapted response decreased markedly (> 40%). Thus neurons increased their rate of adaptation. The afterhyperpolarizations following pulse-evoked responses increased in duration and amplitude with depression. There were input resistance (Rin) reductions at depolarized membrane potentials and during pulses. However, Rin reductions were considerably smaller or altogether absent late during interpulse intervals. Sub-threshold current stimuli were ineffective, indicating that spike activity was necessary to elicit depression. 4. Depression was 1) insensitive to the toxin omega-Agatoxin-IVA (omega-Aga-IVA; 0.5 microM), which blocked synaptic transmission, revealing a key involvement of intrinsic properties and little if any synaptic participation; 2) insensitive to 4-aminopyrydine (2.00-4.00 mM), which greatly enhanced excitatory and inhibitory synaptic efficacy, again suggesting little synaptic involvement and a principal postsynaptic participation, and no participation of the K(+)-mediated currents IA and ID; 3) abolished by carbamalcholine (5.0-20.0 microM)- an effect blocked by atropine (1.0-10.0 microM)- and reduced by Ca(2+)-free solutions, and by intracellular injection of the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), suggesting that Ca(2+)-dependent K(+)-mediated currents are key factors, with a less important participation of the K(+)-mediated IM current. 5. We conclude that depression was due to activity-dependent modifications in intrinsic properties, with little if any synaptic participation. Depression may be functionally significant because it was induced by potentially natural stimulations. A model is proposed that accounts for the main traits of depression. In the model, depression was induced by a gradual decline of the speed at which Ca2+ was buffered intracellularly; an increase in the IK(Ca)S activation rate constant also simulated depression.     

Active summation of excitatory postsynaptic potentials in hippocampal CA3 pyramidal neurons

The manner in which the thousands of synaptic inputs received by a pyramidal neuron are summed is critical both to our understanding of the computations that may be performed by single neurons and of the codes used by neurons to transmit information. Recent work on pyramidal cell dendrites has shown that subthreshold synaptic inputs are modulated by voltage-dependent channels, raising the possibility that summation of synaptic responses is influenced by the active properties of dendrites. Here, we use somatic and dendritic whole-cell recordings to show that pyramidal cells in hippocampal area CA3 sum distal and proximal excitatory postsynaptic potentials sublinearly and actively, that the degree of nonlinearity depends on the magnitude and timing of the excitatory postsynaptic potentials, and that blockade of transient potassium channels linearizes summation. Nonlinear summation of synaptic inputs could have important implications for the computations performed by single neurons and also for the role of the mossy fiber and perforant path inputs to hippocampal area CA3.     

Evidence from simultaneous intracellular recordings in rat hippocampal slices that area CA3 pyramidal cells innervate dentate hilar mossy cells

1. Simultaneous intracellular recordings of area CA3 pyramidal cells and dentate hilar "mossy" cells were made in rat hippocampal slices to test the hypothesis that area CA3 pyramidal cells excite mossy cells monosynaptically. Mossy cells and pyramidal cells were differentiated by location and electrophysiological characteristics. When cells were impaled near the border of area CA3 and the hilus, their identity was confirmed morphologically after injection of the marker Neurobiotin. 2. Evidence for monosynaptic excitation of a mossy cell by a pyramidal cell was obtained in 7 of 481 (1.4%) paired recordings. In these cases, a pyramidal cell action potential was followed immediately by a 0.40 to 6.75 (mean, 2.26) mV depolarization in the simultaneously recorded mossy cell (mossy cell membrane potentials, -60 to -70 mV). Given that pyramidal cells used an excitatory amino acid as a neurotransmitter (Cotman and Nadler 1987; Ottersen and Storm-Mathisen 1987) and recordings were made in the presence of the GABAA receptor antagonist bicuculline (25 microM), it is likely that the depolarizations were unitary excitatory postsynaptic potentials (EPSPs). 3. Unitary EPSPs of mossy cells were prone to apparent "failure." The probability of failure was extremely high (up to 0.72; mean = 0.48) if the effects of all presynaptic action potentials were examined, including action potentials triggered inadvertently during other spontaneous EPSPs of the mossy cell. Probability of failure was relatively low (as low as 0; mean = 0.24) if action potentials that occurred during spontaneous activity of the mossy cell were excluded. These data suggest that unitary EPSPs produced by pyramidal cells are strongly affected by concurrent synaptic inputs to the mossy cell. 4. Unitary EPSPs were not clearly affected by manipulation of the mossy cell's membrane potential. This is consistent with the recent report that area CA3 pyramidal cells innervate distal dendrites of mossy cells (Kunkel et al. 1993). Such a distal location also may contribute to the high incidence of apparent failures. 5. Characteristics of unitary EPSPs generated by pyramidal cells were compared with the properties of the unitary EPSPs produced by granule cells. In two slices, pyramidal cell and granule cell inputs to the same mossy cell were compared. In other slices, inputs to different mossy cells were compared. In all experiments, unitary EPSPs produced by granule cells were larger in amplitude but similar in time course to unitary EPSPs produced by pyramidal cells. Probability of failure was lower and paired-pulse facilitation more common among EPSPs triggered by granule cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

Dendritic hyperpolarization-activated currents modify the integrative properties of hippocampal CA1 pyramidal neurons

Step hyperpolarizations evoked slowly activating, noninactivating, and slowly deactivating inward currents from membrane patches recorded in the cell-attached patch configuration from the soma and apical dendrites of hippocampal CA1 pyramidal neurons. The density of these hyperpolarization-activated currents (Ih) increased over sixfold from soma to distal dendrites. Activation curves demonstrate that a significant fraction of Ih channels is active near rest and that the range is hyperpolarized relatively more in the distal dendrites. Ih activation and deactivation kinetics are voltage-and temperature-dependent, with time constants of activation and deactivation decreasing with hyperpolarization and depolarization, respectively. Ih demonstrated a mixed Na+-K+ conductance and was sensitive to low concentrations of external CsCl. Dual whole-cell recordings revealed regional differences in input resistance (Rin) and membrane polarization rates (taumem) across the somatodendritic axis that are attributable to the spatial gradient of Ih channels. As a result of these membrane effects the propagation of subthreshold voltage transients is directionally specific. The elevated dendritic Ih density decreases EPSP amplitude and duration and reduces the time window over which temporal summation takes place. The backpropagation of action potentials into the dendritic arborization was impacted only slightly by dendritic Ih, with the most consistent effect being a decrease in dendritic action potential duration and an increase in afterhyperpolarization. Overall, Ih acts to dampen dendritic excitability, but its largest impact is on the subthreshold range of membrane potentials where the integration of inhibitory and excitatory synaptic inputs takes place.     

Dendritic properties of hippocampal CA1 pyramidal neurons in the rat: intracellular staining in vivo and in vitro

Dendritic morphology and passive cable properties determine many aspects of synaptic integration in complex neurons, together with voltage-dependent membrane conductances. We investigated dendritic properties of CA1 pyramidal neurons intracellularly labeled during in vivo and in vitro physiologic recordings, by using similar intracellular staining and three-dimensional reconstruction techniques. Total dendritic length of the in vivo neurons was similar to that of the in vitro cells. After correction for shrinkage, cell extent in three-dimensional representation was not different between the two groups. Both in vivo and in vitro neurons demonstrated a variable degree of symmetry, with some neurons showing more cylindrical symmetry around the main apical axis, whereas other neurons were more elliptical, with the variation likely due to preparation and preservation conditions. Branch order analysis revealed no difference in the number of branch orders or dendritic complexity. Passive conduction of dendritic signals to the soma in these neurons shows considerable attenuation, particularly with higher frequency signals (such as synaptic potentials compared with steady-state signals), despite a relatively short electrotonic length. Essential aspects of morphometric appearance and complex dendritic integration critical to CA1 pyramidal cell functioning are preserved across neurons defined from the two different hippocampal preparations used in this study.     

Spontaneous excitatory postsynaptic currents in hippocampal CA1 pyramidal neurons of the gerbil after transient ischemia

The changes in the spontaneous excitatory postsynaptic currents (sEPSCs) after transient cerebral ischemia were studied using whole-cell recording from CA1 pyramidal neurons in the gerbil. In neurons recorded 1-2 days after ischemia, sEPSCs had a slowed time course with the decay time constant fitted by a single exponential and it progressively increased after ischemia. Frequency and amplitude distribution of sEPSCs in ischemic neurons were not significantly different from those in the control neurons. The results support the view that abnormal non-N-methyl-D-aspartic acid currents originate at the degenerated postsynaptic site, unrelated to the presynaptic releasing mechanisms.     

Degeneration of hippocampal CA3 pyramidal cells induced by intraventricular kainic acid

Degeneration of hippocampal CA3 pyramidal cells was investigated by light and electron microscopy after intraventricular injection of the potent convulsant, kainic acid. Electron microscopy revealed evidence of pyramidal cell degeneration within one hour. The earliest degenerative changes were confined to the cell body and proximal dendritic shafts. These included an increased incidence of lysosomal structures, deformation of the perikaryal and nuclear outlines, some increase in background electron density, and dilation of the cisternae of the endoplasmic reticulum accompanied by detachment of polyribosomes. Within the next few hours the pyramidal cells atrophied and became electron dense. Then these cells became electron lucent once more as ribosomes disappeared and their membranes and organelles broke up and disintegrated. Light microscopic changes correlated with these ultrastructural observations. The dendritic spines and the initial portion of the dendritic shaft became electron dense within four hours and degenerated rapidly, whereas the intermediate segment of the dendrites swelled moderately and became more electron lucent. No degenerative changes were evident in pyramidal cell axons and boutons until one day after kainic acid treatment. Less than one hour after kainic acid administration, astrocytes in the CA3 area swelled, initially in the vicinity of the cell body and mossy fiber layers. It is suggested that the paroxysmal discharges initiated in CA3 pyramidal cells by kainic acid served as the stimulus for this response. Phagocytosis commenced between one and three days after kainic acid administration, but remained incomplete at survival times of 6-8 weeks. Astrocytes, microglia, and probably oligodendroglia phagocytized the degenerating material. These results point to the pyramidal cell body and possibly also the dendritic spines as primary targets of kainic acid neurotoxicity. In conjunction with other data, they support the view that lesions made by intraventricular kainic acid can serve as models of epileptic brain damage.     

Cyclooxygenase-2 inhibition prevents delayed death of CA1 hippocampal neurons following global ischemia

The inducible isoform of the enzyme cyclooxygenase-2 (COX2) is an immediate early gene induced by synaptic activity in the brain. COX2 activity is an important mediator of inflammation, but it is not known whether COX2 activity is pathogenic in brain. To study the role of COX2 activity in ischemic injury in brain, expression of COX2 mRNA and protein and the effect of treatment with a COX2 inhibitor on neuronal survival in a rat model of global ischemia were determined. Expression of both COX2 mRNA and protein was increased after ischemia in CA1 hippocampal neurons before their death. There was increased survival of CA1 neurons in rats treated with the COX2-selective inhibitor SC58125 [1-[(4-methylsulfonyl) phenyl]-3-trifluoro-methyl-5-[(4-fluoro)phenyl] pyrazole] before or after global ischemia compared with vehicle controls. Furthermore, hippocampal prostaglandin E2 concentrations 24 h after global ischemia were decreased in drug-treated animals compared with vehicle-treated controls. These results suggest that COX2 activity contributes to CA1 neuronal death after global ischemia.     

Duration of neurofibrillary changes in the hippocampal pyramidal neurons

The clinical course of grave forms of leptospirosis presents with disorders in the fluid and electrolyte balance and acid-base condition (ABC) which fact necessitates taking prompt action for the condition to be corrected. Correction of disorders in the fluid and electrolyte balance involves employment of glucose and salt solutions, dextrans, and in most severe cases albumin drugs under control of hematocrit values, plasma osmolarity, and 24-h diuresis monitoring. Correction of disorders in the ABC is primarily aimed at alleviating the metabolic acidosis through detoxication by applying specific therapy together with oral and parenteral administration of sodium hydrogen carbonate.     

Multiple actions of methohexital on hippocampal CA1 and cortical neurons of rat brain slices

To explore the mechanism by which methohexital (MTH) activates epileptiform activity in patients with epilepsy, we examined the effects of MTH on hippocampal CA1 and neocortical neurons via extracellular and whole-cell patch-clamp recordings in rat brain slices. Perfusion of slices with 10 to 100 microM MTH caused no significant change in glutamatergic transmission in the hippocampal CA1 region, but enhanced gamma-aminobutyric acid (GABA)A-mediated inhibitory postsynaptic currents and induced spontaneous inhibitory postsynaptic currents in neocortical and hippocampal CA1 neurons. In addition, MTH induced a tonic, bicuculline-sensitive hyperpolarization in association with increases in membrane conductance, suggesting a direct stimulation of GABAA receptors by MTH. Spontaneous epileptiform activity was not observed in the neocortex and hippocampus after exposure of slices to MTH, neither in the standard in vitro condition nor in the presence of 4-aminopyridine, which promotes rhythmic synaptic activities. We suggest that the activation of epileptiform activity in vivo by MTH may result from increased neuronal synchrony via the potentiation of GABAA-mediated synaptic inhibition.     

Comparative antagonism of kainate-activated kainate and AMPA receptors in hippocampal neurons

Native kainate receptors expressed by cultured hippocampal cells were studied in the whole-cell configuration of the patch-clamp technique by using a fast perfusion system. About 80% of the neurons expressed kainate receptors independently of the time in culture (0-4 days), which coincided with the number of cells immunoreactive for a monoclonal antibody against the GluR5/6/7 subunits. Three types of cells were considered: neurons in which the rapid application of kainate induced a rapidly desensitizing current, cells in which kainate induced a more slowly rising, non-desensitizing, response and those in which a mixture of both responses was apparent. Steady responses induced by 300 microM kainate were inhibited by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) in a dose-dependent manner (IC50 = 0.92 microM). CNQX was less potent in blocking transient kainate-induced responses (IC50 = 6.1 microM). Responses to kainate, whether steady or transient, were also inhibited by NS102, showing poor selectivity for the transient response (IC50 = 4.1 and 2.2 microM respectively). The new alpha-amino-3-hydroxy-5-methyl-4-isoxazole (AMPA) receptor antagonist NS394 was very potent in inhibiting steady kainate-induced currents (IC50 = 0.45 microM), but was even more effective in preventing peak responses (IC50 = 0.13 microM). In contrast, cyclothiazide did not affect transient kainate-induced responses but did potentiate current induced by activation of AMPA receptors by AMPA or kainate. These results demonstrate the lack of complete selectivity amongst some available competitive antagonists for AMPA and kainate receptors, and indicate that kainate receptors expressed by hippocampal cells lack the cyclothiazide modulatory site present at AMPA receptors. In addition, the present data support the idea that low-affinity kainate binding sites in the brain correspond to receptor channels selectively activated by kainate.     

Postischemic enhancements of N-methyl-D-aspartic acid (NMDA) and non-NMDA receptor-mediated responses in hippocampal CA1 pyramidal neurons

Glutamate receptor-mediated responses were investigated by using a whole-cell recording and an intracellular calcium ion ([Ca2+]i) imaging in gerbil postischemic hippocampal slices prepared at 1, 3, 6, 9, 12, and 24 hours after 5-minute ischemia. Bath application of N-methyl-D-aspartic acid (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), and kainate showed that NMDA-, AMPA- and kainate-induced currents were enhanced in postischemic CA1 pyramidal neurons at 1 to 12 hours after 5-minute ischemia. NMDA and non-NMDA receptor-mediated excitatory postsynaptic currents (EPSC) were examined in postischemic CA1 pyramidal neurons at 3 hours after 5-minute ischemia to confirm whether synaptic responses are enhanced in the postischemic CA1 pyramidal neurons. The amplitudes of NMDA- and non-NMDA-receptor-mediated EPSC were enhanced in the postischemic CA1 pyramidal neurons. NMDA-, AMPA-, and kainate-induced [Ca2+]i elevations were also examined to determine whether the enhancement of currents is accompanied by the enhancement of [Ca2+]i elevation. The enhancements of NMDA-, AMPA-, and kainate-induced [Ca2+]i elevations were shown in the postischemic CA1. These results indicate that NMDA and non-NMDA receptor-mediated responses are persistently enhanced in the CA1 pyramidal neurons 1 to 12 hours after transient ischemia, and suggest that the enhancement of glutamate receptor-mediated responses may act as one of crucial factors in the pathologic mechanism responsible for leading postischemic CA1 pyramidal neurons to irreversible neuronal injury.     

Ethanol inhibition of AMPA and kainate receptor-mediated depolarizations of hippocampal area CA1

Longitudinal hippocampal slices were prepared from adult female rats. The excitatory amino acids, alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and kainic acid, were applied to area CA1, and the resulting depolarizations were measured using the grease-gap electrophysiological technique. Agonist dose-response curves were generated in the presence and absence of various concentrations of ethanol. Ethanol (25-200 mM) significantly attenuated the depolarizations that were produced by each agonist. In addition, we found that ethanol potently antagonized kainate-induced depolarizations across the agonist concentration-response curve, whereas it significantly suppressed only AMPA responses that were induced with moderate-to-high agonist concentrations. These results indicate that ethanol has potent antagonist actions against non-N-methyl-D-aspartate (NMDA) excitatory amino acid-induced neuronal depolarizations in hippocampal area CA1. Moreover, the relative potency of ethanol depends on the specific excitatory agonist tested and the concentration of that agonist. This suggests that, in addition to the known effects of ethanol on NMDA receptor-mediated activity, it may also potently attenuate ongoing "fast" glutamatergic synaptic activity in the hippocampus.     

Interleukin-1beta does not increase synaptic inhibition in hippocampal CA3 pyramidal and dentate gyrus granule cells of the rat in vitro

Effects of interleukin-1beta (bath-applied; 500 pM) on rat hippocampal CA3 pyramidal and dentate granule cells were studied using intracellular microelectrode recording in vitro. In both cell types membrane input resistance, resting membrane potential and action potential amplitude remained stable throughout. No change was seen in postsynaptic potentials in granule cells. After blocking excitatory synaptic transmission in CA3 pyramids interleukin-1beta was found to consistently decrease synaptic inhibition by about 30%.     

Differentiation of neurons and synapses of mossy fibers in transplants of fascia dentata developing in the rat neocortex]

Lipopolysaccharide is an inflammatory agent and interleukin-1 is a cytokine. Their pro-inflammatory effects may be mediated by prostanoids produced by inducible cyclooxygenase-2. The aim of this study was to determine the prostanoids produced by lipopolysaccharide and interleukin-1 stimulated enterocytes through the cyclooxygenase-1 and 2 pathways. Cultured enterocytes were stimulated with lipopolysaccharide or interleukin-1beta with and without cyclooxygenase inhibitors. Low concentrations of indomethacin and valerylsalicylic acid (VSA) were evaluated as cyclooxygenase-1 inhibitors and their effects compared with the effects of a specific cyclooxygenase-2 inhibitor, SC-58125. Prostaglandin E2, 6-keto prostaglandin F1alpha, prostaglandin D2 and leukotriene B4 levels were determined by radioimmunoassay. Immunoblot analysis using isoform-specific antibodies showed that the inducible cyclooxygenase enzyme (COX-2) was expressed by 4 h in LPS and IL-1beta treated cells while the constitutive COX-1 remained unaltered in its expression. Interleukin-1beta and lipopolysaccharide stimulated the formation of all prostanoids compared with untreated cells, but failed to stimulate leukotriene B4. Indomethacin at 20 microM concentration, and VSA inhibited lipopolysaccharide and interleukin 1beta stimulated prostaglandin E2, but not 6-keto prostaglandin F1alpha formation. SC-58125 inhibited lipopolysaccharide and interleukin-1beta stimulated 6-keto prostaglandin F1alpha but not prostaglandin E2 release. The specific cyclooxygenase-2 inhibitor also inhibited lipopolysaccharide produced prostaglandin D2 but not interleukin-1beta stimulated prostaglandin D2. While SC-58125 inhibited basal 6-keto prostaglandin-F1alpha formation it significantly increased basal prostaglandin E2 and prostaglandin D2 formation. As SC-58125 inhibited lipopolysaccharide and interleukin-1beta induced 6-keto prostaglandin F1alpha production but not prostaglandin E2 production, it suggests that these agents stimulate prostacyclin production through a cyclooxygenase-2 mediated mechanism and prostaglandin E2 production occurs through a cyclooxygenase-1 mediated mechanism. Prostaglandin D2 production appeared to be variably produced by cyclooxygenase-1 or cyclooxygenase-2, depending on the stimulus.     

Influence of the dentate fascia on the sensory responses of neurons in hippocampal field CA3

In experiments with unanaesthetized rabbits the influences of electric stimulation of the dentate fascia (DF) on the extracellularly recorded spontaneous and evoked activity of the CA3 neurones were investigated. Stimulation of a fixed locus in the DF during recording in the CA3 by a microelectrode, shifted along the longitudinal axis of the hippocampus, supported the notion of the topical, "segmental" organization of connections between the DF and CA3. A relatively narrow "active zone" (approximately 700 nm) appeared in the CA3 during the threshold DF stimulation: it was bordered by zones with predominantly inhibitory responses to stimulation. The CA3 neurones in the "active zone" rapidly lost their reactivity to sensory stimuli. In the "inhibitory" and "zero" zones the normal level of reactivity to sensory stimuli was preserved.     

Downregulation of transient K+ channels in dendrites of hippocampal CA1 pyramidal neurons by activation of PKA and PKC

A monoclonal antibody (MoAb) specific for the human P2X7 receptor was generated in mice. As assessed by flow cytometry, the MoAb labeled human blood-derived macrophage cells natively expressing P2X7 receptors and cells transfected with human P2X7 but not other P2X receptor types. The MoAb was used to immunoprecipitate the human P2X7 receptor protein, and in immunohistochemical studies on human lymphoid tissue, P2X7 receptor labeling was observed within discrete areas of the marginal zone of human tonsil sections. The antibody also acted as a selective antagonist of human P2X7 receptors in several functional studies. Thus, whole cell currents, elicited by the brief application of 2',3'-(4-benzoyl)-benzoyl-ATP in cells expressing human P2X7, were reduced in amplitude by the presence of the MoAb. Furthermore, preincubation of human monocytic THP-1 cells with the MoAb antagonized the ability of P2X7 agonists to induce the release of interleukin-1beta.