1. Intracellular current-clamp recordings were made from neurons of the basolateral nucleus of the amygdala (BLA) of the rat in the in vitro slice preparation. Neurons were identified morphologically after intracellular injection of biocytin, and the electrophysiological properties and morphological characteristics were correlated. 2. Three distinct morphological subtypes were identified: Class I pyramidal neurons, Class I stellate neurons, and Class II neurons. Each morphological subtype could also be distinguished according to its characteristic electrophysiological properties. 3. Class I pyramidal neurons typically had pyramidal perikarya (cross-sectional area = 245 microns2) with spine-laden apical and basal dendrites. The axon originated from the largest basal dendrite and produced several collaterals that ramified throughout the dendritic arborization of the parent cell. These neurons were characterized electrophysiologically by their higher input resistance (65.6 M omega), long time constant of membrane charging tau 0 (27.8 ms), long duration action potential (half-width = 0.85 ms), and regular firing pattern [1st interspike interval ISI) = 91 ms]. 4. Class I stellate neurons differed morphologically from Class I pyramidal neurons only in the size (cross sectional area = 330 microns 2) and stellate appearance of their perikarya. These neurons had characteristic lower input resistance (40.1 M omega), shorter time constant of membrane charging tau 0 (14.5 ms), shorter duration action potential (half-width = 0.7 ms), and a burst firing pattern (1st ISI = 6.0 ms), all of which were statistically different from Class I pyramidal neurons. 5. Class II neurons were multipolar (cross sectional area = 235 microns 2) and were distinguishable from Class I neurons by the almost complete absence of dendritic spines. Class II neurons were characterized electrophysiologically by a midrange input resistance (58 M omega), intermediate time constant of membrane charging tau 0 (19 ms), intermediate action-potential duration (half-width = 0.77 ms), and a burst firing pattern (1st ISI = 6.0 ms). In contrast to Class I neurons, action-potential firing of Class II neurons did not accommodate in response to prolonged depolarizing current injection. 6. In conclusion, BLA neurons may be characterized by their specific electrophysiological properties as well as by their morphological traits. Therefore, permitting assessment of signal transduction in identified populations of neurons within this nucleus.

1. Intracellular recording techniques were used to characterize synaptic inhibitory postsynaptic potentials (IPSPs) recorded from neurons of the basolateral nucleus of the amygdala (BLA). Bipolar electrodes positioned in the stria terminalis (ST) or lateral amygdala (LA) were used to evoke synaptic responses at a frequency of 0.25 Hz. 2. Two synaptic waveforms having IPSP components could be evoked by electrical stimulation of either pathway: a biphasic, excitatory postsynaptic potential (EPSP), fast-IPSP (f-IPSP) waveform, and a multiphasic, EPSP, f-IPSP, and subsequent slow-IPSP (s-IPSP) waveform. Expression of either waveform was dependent on the site of stimulation. ST stimulation evoked a similar number of biphasic (45%) and multiphasic (50%) synaptic responses. In contrast, stimulation of the LA pathway evoked mainly (80%) multiphasic synaptic responses. 3. Both the f- and s-IPSP elicited by ST stimulation could be reduced in amplitude in the presence of the glutamatergic, N-methyl-D-aspartate (NMDA) antagonist, (DL)-2-amino-5-phosphonovaleric acid (APV, 50 microM), and were abolished by the glutamatergic, non-NMDA antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 microM). In contrast, a CNQX-resistant f-IPSP was evoked with LA stimulation and abolished by subsequent addition of bicuculline methiodide (BMI), a gamma-aminobutyric acid (GABAA) receptor antagonist, suggesting direct inhibition of BLA neurons by GABAergic LA interneurons. The sensitivity of the s-IPSPs and the f-IPSPs to glutamatergic antagonists suggests the presence of feed-forward inhibition onto BLA neurons. 4. The f-IPSP possessed characteristics of potentials mediated by GABAA receptors linked to Cl- channels, namely, a reversal potential of -70 mV, a decrease in membrane resistance (13.5 M omega) recorded at -60 mV, a block by BMI, and potentiation by sodium pentobarbital (NaPB). 5. The s-IPSP was associated with a resistance decrease of 4.5 M omega, a reversal potential of -95 mV, and was reversibly depressed (approximately 66%) by 2-hydroxy-saclofen (100 microM), suggesting activation of GABAB receptors. 6. The large resistance change associated with the f-IPSP, its temporal overlap with evoked EPSPs, and the development of both spontaneous and evoked burst firing in the presence of BMI suggests that the f-IPSP determines the primary state of excitability in BLA neurons.(ABSTRACT TRUNCATED AT 400 WORDS)

A RP-HPLC method was developed for the assessment of caffeine and its metabolites in urine and was used for the evaluation of the CYP1A2, CYP2A6, xanthine oxidase (XO) and N-acetyl-transferase-2 (NAT-2) in vivo activities in 44 Greek volunteers (21 men, 23 women). Spot urine samples were analyzed 6 h after 200 mg caffeine consumption, following a 30 h methylxantine-free diet. The major urinary caffeine metabolites are 1-methyluric acid (1U), 5-acetylamino-6-formylamino-3-methyluracil (AFMU), 1-methylxanthine (1X), 1,7-dimethyluric acid (17U) and 1,7-dimethylxanthine (17X). CYP1A2, CYP2A6, XO and NAT-2 activities were estimated from the metabolic ratios (AFMU + 1U + 1X)/17U, 17U/17X, 1U/(1X + 1U) and AFMU/(AFMU + 1U + 1X), respectively. Metabolites and internal standard were extracted with chloroform/isopropanol (85:15, v/v) and separated on a C 18 column by an isocratic HPLC system using a two-step elution with manual switch from solvent A (0.1% acetic acid-methanol-acetonitrile, 92:4:5 v/v) to solvent B (0.1% acetic acid-methanol, 60:40, v/v), and detected at 280 nm. The method exhibited adequate metabolite separation (resolution factors >1.48), accuracy (94.1-106.3%) and intraday and interday precision <8.02 and <8.78%, respectively (n = 6). Smoking affected only CYP1A2, whereas gender had no effect in any enzyme activity. NAT-2 exhibited bimodal distribution, 63.6% of volunteers being slow acetylators. The developed RP-HPLC method was fully validated and successfully applied for the evaluation of CYP1A2, CYP2A6, XO and NAT-2 activities. Copyright

The action of ethosuximide (ETX) on Na+, K+, and Ca2+ currents and on tonic and burst-firing patterns was investigated in rat and cat thalamic neurons in vitro by using patch and sharp microelectrode recordings. In thalamocortical (TC) neurons of the rat dorsal lateral geniculate nucleus (LGN), ETX (0.75-1 mM) decreased the noninactivating Na+ current, INaP, by 60% but had no effect on the transient Na+ current. In TC neurons of the rat and cat LGN, the whole-cell transient outward current was not affected by ETX (up to 1 mM), but the sustained outward current was decreased by 39% at 20 mV in the presence of ETX (0.25-0.5 mM): this reduction was not observed in a low Ca2+ (0.5 mM) and high Mg2+ (8 mM) medium or in the presence of Ni2+ (1 mM) and Cd2+ (100 microM). In addition, ETX (up to 1 mM) had no effect on the low-threshold Ca2+ current, IT, of TC neurons of the rat ventrobasal (VB) thalamus and LGN and in neurons of the rat nucleus reticularis thalami nor on the high-threshold Ca2+ current in TC neurons of the rat LGN. Sharp microelectrode recordings in TC neurons of the rat and cat LGN and VB showed that ETX did not change the resting membrane potential but increased the apparent input resistance at potentials greater than -60 mV, resulting in an increase in tonic firing. In contrast, ETX decreased the number of action potentials in the burst evoked by a low-threshold Ca2+ potential. The frequency of the remaining action potentials in a burst also was decreased, whereas the latency of the first action potential was increased. Similar effects were observed on the burst firing evoked during intrinsic delta oscillations. These results indicate an action of ETX on INaP and on the Ca2+-activated K+ current, which explains the decrease in burst firing and the increase in tonic firing, and, together with the lack of action on low- and high-threshold Ca2+ currents, the results cast doubts on the hypothesis that a reduction of IT in thalamic neurons underlies the therapeutic action of this anti-absence medicine.

1. Intracellular current-clamp recordings obtained from neurons of the basolateral nucleus of the amygdala (BLA) were used to characterize postsynaptic potentials elicited through stimulation of the stria terminalis (ST) or the lateral amygdala (LA). The contribution of glutamatergic receptor subtypes to excitatory postsynaptic potentials (EPSPs) were analyzed by the use of the non N-methyl-D-aspartate (non-NMDA) antagonist, 6-cyano-7-nitro-quinoxaline-2,3-dione (CNQX), and the NMDA antagonist, (DL)-2-amino-5-phosphonovaleric acid (APV). 2. Basic membrane properties of BLA neurons determined from membrane responses to transient current injection showed that at the mean resting membrane potential (RMP; -67.2 mV) the input resistance (RN) and time constant for membrane charging (tau) were near maximal, and that both values were reduced with membrane hyperpolarization, suggesting an intrinsic regulation of synaptic efficacy. 3. Responses to stimulation of the ST or LA consisted of an EPSP followed by either a fast inhibitory postsynaptic potential (f-IPSP) only, or by a fast- and subsequent slow-IPSP (s-IPSP). The EPSP was graded in nature, increasing in amplitude with increased stimulus intensity, and with membrane hyperpolarization after DC current injection. Spontaneous EPSPs were also observed either as discrete events or as EPSP/IPSP waveforms. 4. In physiological Mg2+ concentrations (1.2 mM), at the mean RMP, the EPSP consisted of dual, fast and slow, glutamatergic components. The fast-EPSP (f-EPSP) possessed characteristics of kainate/quisqualate receptor activation, namely, the EPSP increased in amplitude with membrane hyperpolarization, was insensitive to the NMDA receptor antagonist, APV (50 microM), and was blocked by the non-NMDA receptor antagonist, CNQX (10 microM). In contrast, the slow-EPSP (s-EPSP) decreased in amplitude with membrane hyperpolarization, was insensitive to CNQX (10 microM), and was blocked by APV (50 microM), indicating mediation by NMDA receptor activation. 5. In the presence of CNQX (10 microM), ST stimulation evoked an APV-sensitive s-EPSP. In contrast, LA stimulation evoked a f-IPSP, which when blocked by subsequent addition of bicuculline methiodide (BMI; 30 microM) revealed a temporally overlapping APV-sensitive s-EPSP. These data suggest that EPSP amplitude and duration are determined, in part, by the shunting of membrane conductance caused by a concomitant IPSP. 6. Superfusion of either CNQX or APV in BLA neurons caused membrane hyperpolarization and blockade of spontaneous EPSPs and IPSPs, suggesting that these compounds may act to block tonic excitatory amino acid (EAA) release within the nucleus, and that a degree of feed-forward inhibition occurs within the nucleus.(ABSTRACT TRUNCATED AT 400 WORDS)