Biomedical subjects

Ricardo M Leão

Publications and source records attributed to Ricardo M Leão.

5 recordsLinked to original sources

PubMed2006

Urethane inhibits the GABAergic neurotransmission in the nucleus of the solitary tract of rat brain stem slices.

Because urethane is a widely used anesthetic in animal experimentation, in the present study, we evaluated its effects on neurons of the nucleus of the solitary tract (NTS) in brain stem slices from young rats (25-30 days old). Using the whole cell configuration of the patch-clamp technique, spontaneous postsynaptic currents (sPSCs) and evoked excitatory postsynaptic currents (eEPSCs) were recorded. Urethane (20 mM) decreased by approximately 60% the frequency of GABAergic sPSCs (1.0 +/- 0.2 vs. 0.4 +/- 0.1 Hz) but did not change the frequency, amplitude, or half-width of glutamatergic events or TTX-resistant inhibitory sPSCs [miniature inhibitory postsynaptic currents (IPSCs)]. Miniature IPSCs were measured in the presence of urethane plus 1 mM diazepam (1 mM), and no changes were seen in their amplitude. This suggests that the GABA concentration in the NTS synapses is set at saturating level. We also evaluated the effect of urethane on eEPSCs, and no significant change was observed in the amplitude of N-methyl-d-aspartate [NMDA; 44.2 +/- 11.5 vs. 37.6 +/- 10.6 pA (holding potential = 40 mV)] and non-NMDA currents [204.4 +/- 35.5 vs. 196.6 +/- 31.2 pA (holding potential = -70 mV)]. Current-clamp experiments showed that urethane did not alter the action potential characteristics and passive membrane properties. These data suggest that urethane has an inhibitory effect on GABAergic neurons in the NTS but does not change the spontaneous or evoked excitatory responses.

Anesthetics, Intravenous↗

PubMed2005

Presynaptic Na+ channels: locus, development, and recovery from inactivation at a high-fidelity synapse.

Na+ channel recovery from inactivation limits the maximal rate of neuronal firing. However, the properties of presynaptic Na+ channels are not well established because of the small size of most CNS boutons. Here we study the Na+ currents of the rat calyx of Held terminal and compare them with those of postsynaptic cells. We find that presynaptic Na+ currents recover from inactivation with a fast, single-exponential time constant (24 degrees C, tau of 1.4-1.8 ms; 35 degrees C, tau of 0.5 ms), and their inactivation rate accelerates twofold during development, which may contribute to the shortening of the action potential as the terminal matures. In contrast, recordings from postsynaptic cells in brainstem slices, and acutely dissociated, reveal that their Na+ currents recover from inactivation with a double-exponential time course (tau(fast) of 1.2-1.6 ms; tau(slow) of 80-125 ms; 24 degrees C). Surprisingly, confocal immunofluorescence revealed that Na+ channels are mostly absent from the calyx terminal but are instead highly concentrated in an unusually long (approximately 20-40 microm) unmyelinated axonal heminode. Outside-out patch recordings confirmed this segregation. Expression of Na(v)1.6 alpha-subunit increased during development, whereas the Na(v)1.2alpha-subunit was not present. Serial EM reconstructions also revealed a long pre-calyx heminode, and biophysical modeling showed that exclusion of Na+ channels from the calyx terminal produces an action potential waveform with a shorter half-width. We propose that the high density and polarized locus of Na+ channels on a long heminode are critical design features that allow the mature calyx of Held terminal to fire reliably at frequencies near 1 kHz.

Action Potentials↗

PubMed2004

GABAergic neurons participate in the brain's response to birdsong auditory stimulation.

Birdsong is a learned vocal behaviour that requires intact hearing for its development in juveniles and for its maintenance during adulthood. However, the functional organization of the brain circuits involved in the perceptual processing of song has remained obscure. Here we provide evidence that GABAergic mechanisms are an important component of these circuits and participate in the auditory processing of birdsong. We first cloned a zebra finch homologue of the gene encoding the 65-kDa isoform of glutamic acid decarboxylase (zGAD-65), a specific GABAergic marker, and conducted an expression analysis by in situ hybridization to identify GABAergic cells and to map their distribution throughout auditory telencephalic areas. The results showed that field L2, the caudomedial nidopallium (NCM) and the caudomedial mesopallium (CMM) contain a high number of GABAergic cells. Using patch-clamp brain slice recordings, we found abundant GABAergic mIPSCs in NCM. Pharmacological antagonism of mIPSCs induced large EPSC bursts, suggesting that tonic inhibition helps to stabilize NCM against runaway excitation via activation of GABA-A receptors. Next, using double fluorescence in situ hybridization and double immunocytochemical labelling, we demonstrated that large numbers of GABAergic cells in NCM and CMM show inducible expression of the transcriptional regulator ZENK in response to song auditory stimulation. These data provide direct evidence that GABAergic neurons in auditory brain regions are activated by song stimulation. Altogether, our results suggest that GABAergic mechanisms participate in auditory processing and perception, and might contribute to the memorization of birdsong.

Acoustic Stimulation↗

PubMed2002

Optimizing synaptic architecture and efficiency for high-frequency transmission.

Bursts of neuronal activity are transmitted more effectively as synapses mature. However, the mechanisms that control synaptic efficiency during development are poorly understood. Here, we study postnatal changes in synaptic ultrastructure and exocytosis in a calyx-type nerve terminal. Vesicle pool size, exocytotic efficiency (amount of exocytosis per Ca influx), Ca current facilitation, and the number of active zones (AZs) increased with age, whereas AZ area, number of docked vesicles per AZ, and release probability decreased with age. These changes led to AZs that are less prone to multivesicular release, resulting in reduced AMPA receptor saturation and desensitization. A greater multiplicity of small AZs with few docked vesicles, a larger pool of releasable vesicles, and a higher efficiency of release thus promote prolonged high-frequency firing in mature synapses.

Action Potentials↗

PubMed2002

Noradrenaline increases high-frequency firing at the calyx of Held synapse during development by inhibiting glutamate release.

The mammalian auditory brain stem receives profuse adrenergic innervation, whose function is poorly understood. Here we investigate, during postnatal development, the effect of noradrenaline (NA) at the calyx of Held synapse in the rat medial nucleus of the trapezoid body (MNTB). We observed that NA inhibits the large glutamatergic EPSC, evoked by afferent fiber stimulation, in a dose-dependent manner. The inhibition was maximal (approximately 48%) at the concentration of 2 microM. It was antagonized by yohimbine and mimicked by the alpha2-adrenergic specific agonist UK14304. Both AMPA and NMDA receptor-mediated EPSCs were inhibited in parallel by NA, suggesting a presynaptic effect. Presynaptic recordings showed that NA inhibits the action potential (AP) generated Ca current by about 20%; however, NA did not significantly affect the presynaptic AP waveform. We thus conclude that the calyx of Held presynaptic terminal expresses alpha2-adrenergic receptors that inhibit its Ca current and thus glutamate release. Noradrenaline was effective in all cells tested from postnatal days 6 to 7 (P6-P7), and thereafter the number of responsive cells diminished, although half of the P14 cells tested still had EPSCs that were inhibited by NA. By contrast, activation by L-2-amino-5-phosphonovaleric acid-sensitive metabotropic glutamate receptors strongly inhibited the EPSCs of all cells tested from P6 to P14. The effect of NA on postsynaptic action potential firing was dependent on the stimulus frequency. At 10 Hz, NA had no effect on firing probability; however, NA helped MNTB cells fire more action potentials during a 100-Hz train of stimuli, even though it did not increase the steady-state depressed EPSC, because it produced a smaller N-methyl-D-aspartate (NMDA) receptor-activated depolarizing plateau. We therefore suggest that the reduction by NA of the first few EPSCs in a train leads to a smaller NMDA depolarizing plateau and thus to increased firing probability at 100 Hz in young synapses. Surprisingly, the inhibition of glutamate release by NA can thus actually increase the excitability of MNTB neurons during early postnatal development.

Action Potentials↗