Involvement of divalent ions in the nitric oxide-induced blockade of N-methyl-D-aspartate receptors in cerebellar granule cells.

We have previously shown that nitric oxide blocks the N-methyl-D-aspartate (NMDA) receptor without affecting the agonist binding site. We now report that in cerebellar granule cells nitric oxide decreases the NMDA channel conductance and open probability, in voltage-dependent and -independent manners, respectively, by acting on an extracellular site different from the redox, glycine, and pH modulatory sites of the receptor-channel complex. This inhibition is not additive with those of Mg2+ and Zn2+.

Get receptive to metabotropic glutamate receptors.

Glutamate activates not only ionotropic glutamate receptors, but also G-protein-coupled receptors, called metabotropic glutamate receptors. Recent studies have revealed that these metabotropic receptors share distinctive structural properties and that they form a subgroup within the heptahelical receptor family. The development of ligands that bind specifically to these receptors has provided a means of characterizing the important roles they play in the tuning of fast synaptic transmission, including the induction of long-term changes in synaptic strength.

Action of argiotoxin636 on N-methyl-D-aspartate channels in cerebellar cells.

We investigated the mode of action of argiotoxin636 on isolated N-methyl-D-aspartate (NMDA) receptor channels in cultured cerebellar granule cells. We found that the toxin blocks NMDA channels by decreasing their opening probability and by inducing a flickering activity, in a voltage-dependent manner. Our results indicate that argiotoxin636 acts as an open-channel blocker and might therefore be a useful tool for studying the structure of glutamate-gated channels.

Molecular, functional, and pharmacological characterization of the metabotropic glutamate receptor type 5 splice variants: comparison with mGluR1.

The main excitatory neurotransmitter in the brain, glutamate (Glu), activates not only receptor-channels, but also receptors coupled to G-protein called metabotropic Glu receptors (mGluRs). Eight genes coding for mGluRs have been characterized to date giving rise to even more proteins due to alternative splicing phenomena. Here we characterized a splice variant of mGluR5, called mGluR5b which contains a 32 amino acid fragment inserted in the cytoplasmic tail, 50 residues after the 7th transmembrane domain.

Differential expression of pituitary adenylate cyclase-activating polypeptide/vasoactive intestinal polypeptide receptor subtypes in clonal pituitary somatotrophs and gonadotrophs.

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) are hypothalamic factors believed to play a role in the regulation of anterior pituitary cell function. However, little is known about the expression of PACAP/VIP receptor (PVR) subtypes in such cells. Three PVR subtypes have recently been cloned: the PACAP-selective PVR1, and PVR2 and PVR3, which exhibit similar affinities for PACAP and VIP.

Homologous desensitization of 5-hydroxytryptamine4 receptors in rat esophagus: functional and second messenger studies.

We have studied agonist-induced desensitization of 5-hydroxytryptamine (5-HT4) receptor-mediated relaxation and 5-HT4 receptor-mediated increases in cAMP in rat esophageal tunica muscularis mucosae. In both cases, the desensitization time course was biphasic. The first phase was very rapid because more than 50% of desensitization was obtained after a 5-min incubation period with 10 microM of 5-HT.

[The role of nitric oxide and superoxides in the neurotoxicity of glutamate].

Glutamate is the major neurotransmitter of the mammalian brain. Stimulation of glutamate receptors, especially the subgroup of NMDA receptors, induces nitric oxide and arachidonic acid synthesis in neurons. These agents freely diffuse across membranes and thus can play roles of messengers in particular brain functions.

Epidemiology of interstitial lung disease (ILD) in flanders: registration by pneumologists in 1992-1994. Working group on ILD, VRGT. Vereniging voor Respiratoire Gezondheidszorg en Tuberculosebestrijding.

Worldwide almost no epidemiologic data are available on the prevalence or incidence of interstitial lung diseases (ILD) in the general population. Therefore, a registration programme of ILD-prevalence was organised by the VRGT (Vereniging voor Respiratoire Gezondheidszorg en Tuberculosebestrijding), among about 100 Flemish pneumologists since 1990. Most categories of the classification by Crystal et al.

Facilitatory coupling between a glutamate metabotropic receptor and dihydropyridine-sensitive calcium channels in cultured cerebellar granule cells.

The effect of metabotropic glutamate receptor activation on Ca dihydropyridine (DHP)-sensitive channels recorded in the presence of 1 microM Bay K 8644 was examined on cultured cerebellar granule cells using the patch-clamp technique in the cell-attached configuration. Bath-applied agonist (trans-ACPD, 1S,3R-, and 1R,3S-ACPD isomers, and glutamate or quisqualate in the presence of CPP and CNQX) evoked an increase in Ca channel activity with a variable latency of 8.9 +/- 8.

NADPH diaphorase-positive cells in the brain after status epilepticus.

Using NADPH-diaphorase (NADPH-d) histochemistry, the expression of nitric oxide synthase (NOS) was studied in the rat brain 1 week after kainate-induced status epilepticus. Major changes were observed in the hippocampi of epileptic animals, especially a loss of NADPH-d positive fibres in the periphery of degenerative pyramidal cells, the survival of NOS-containing interneurones in the dentate hilus, a different pattern of NADPH-d staining in lesioned areas, probably corresponding to the expression of inducible NOS by glial cells and an increased staining of the vasculature. These different sources of NO may exert different functions in the epileptic focus.

5-HT6 receptors positively coupled to adenylyl cyclase in striatal neurones in culture.

5-HT receptor positively coupled to adenylyl cyclase in striatal neurones in culture does not correspond to the 5-HT4 receptor. 5-HT induces an increase in cAMP level with an EC50 of 125 nM. 5-HT agonists displayed the following rank order of potencies 5-HT > LSD > 5-MeOT > 5-CT.

Up-regulation in late pregnancy of both Go1 alpha and Go2 alpha isoforms in human myometrium.

The nature of the 39 kDa pertussis toxin substrate previously detected in human pregnant myometrium was investigated. Comparison of membranes from non-pregnant and from 39-40 week pregnant myometrium revealed a higher level at 39 kDa of pertussis toxin ADP-ribosylation and Go alpha immunoreactivity in late pregnancy. Furthermore, quantification of both Go alpha isoforms with specific anti-alpha o1- and alpha o2-antibodies revealed an increase in their expression in late pregnancy.

Inhibitory effects of dihydropyridines on macroscopic K+ currents and on the large-conductance Ca(2+)-activated K+ channel in cultured cerebellar granule cells.

In cultured cerebellar granule cells, we examined the effects of dihydropyridines (DHPs) on K+ currents, using the whole-cell recording configuration of the patch-clamp technique and on Ca(2+)-activated K+ channels ("maxi K+ channels") using outside-out patches. We found that micromolar concentrations of nicardipine, nifedipine, (+) and (-) BAY K 8644, nitrendipine, nisoldipine and (-) nimodipine block 10-60% of macroscopic K+ currents. The most potent of these DHPs was nicardipine and the least potent, (-) BAY K 8644.

Identification of multiple subunits of heterotrimeric G proteins on the membrane of secretory granules in rat prolactin anterior pituitary cells.

The subcellular distribution of multiple subunits of heterotrimeric GTP-binding proteins has been investigated in rat anterior pituitary cells in primary culture, and more precisely in prolactin cells, by immunocytochemistry and subcellular fractionation followed by immunoblotting or ADP ribosylation, using polyclonal affinity-purified antibodies directed against Gi3 alpha, Gs alpha, Go1 alpha, Go2 alpha, and G beta. As expected, all these subunits were detected on the plasma membrane. They were, however, also detected on the membrane of several intracellular compartments involved in the secretory pathway, particularly on the secretory granule membrane.

The metabotropic glutamate receptor types 2/3 inhibit L-type calcium channels via a pertussis toxin-sensitive G-protein in cultured cerebellar granule cells.

Modulation of Ca2+ channels by metabotropic glutamate receptors (mGluRs) was investigated in cerebellar granule cells using the cell-attached configuration of the patch-clamp technique. Experiments were performed in the absence of external Ca2+ and Ba2+ was used as charge carrier. Bath applied glutamate or (1S,3R) trans-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R t-ACPD) inhibited Ca2+ channels activated by depolarizing pulses.

Blockade of nitric oxide synthesis by tyrosine kinase inhibitors in neurones.

In striatal neurones in culture, N-methyl-D-aspartate-(NMDA), kainate-(Kai) and K(+)-dependent cGMP production is entirely mediated via nitric oxide (NO). Low concentrations of lavendustin-A (< or = 0.3 microM), a highly specific tyrosine kinase inhibitor, reduced irreversibly and in a time-dependent manner NMDA-stimulated cGMP production.

Plasticity of NMDA receptor expression during mouse cerebellar granule cell development.

A period of hypersensitivity to N-methyl-D-aspartate (NMDA) has been described during the early development of different types of neuron. Since activation of NMDA receptors can also induce rapid neuron death, the hypersensitivity to NMDA may be tightly controlled. In the present study we show that mouse cerebellar granule neurons become transiently hypersensitive to NMDA between days 10 and 14 after plating in a culture medium containing 30 mM K+.

Mutation of valine residue unique to alpha subunit of Gs abolishes activation.

We recently characterized a decapeptide sequence (residues 367-376) that is important for the membrane association of the activated alpha subunit of Gs. We report here that when this sequence is replaced by the cognate sequence of Gi1 alpha subunit, the chimeric protein (Gsis alpha) still interacts with the membrane but cannot be activated, regardless of the mode of activation. Construction of various chimeras demonstrates that the single replacement of valine 367 by threonine, the cognate residue of Gi1 alpha subunit, fully reproduces the loss of activation.

The PACAP receptor: generation by alternative splicing of functional diversity among G protein-coupled receptors in nerve cells.

Recent molecular characterization of new G protein-coupled receptors (GPCR) draw attention to alternative splicing as a source of structural diversity. After a brief overview of characterized GPCR splice variants, we will describe in more detail the functional properties of the PACAP type I receptor splice variants. Some of these variants are positively coupled to both adenylate cyclase (AC) and phospholipase C (PLC) whereas others do not elicit any stimulation of the PLC or display a qualitatively intermediate phenotype.

5-Hydroxytryptamine1A receptor synthetic peptides. Mechanisms of adenylyl cyclase inhibition.

The 5-hydroxytryptamine1A receptor (5-HT1AR) is a G-protein-coupled receptor negatively coupled to adenylyl cyclase (AC). We have studied the functional domains of 5-HT1AR using synthetic peptides to block or mimic receptor function. The entire second intracellular loop (5-HT1AR-i2) and the carboxyl end of the third intracellular loop (5-HT1AR-i3-C) strongly inhibited forskolin-stimulated AC activity.

Glutamate receptors induce a burst of superoxide via activation of nitric oxide synthase in arginine-depleted neurons.

We have previously shown in cultured cerebellar granule neurons (Lafon-Cazal, M., Pietri, S., Culcasi, M.

Pharmacological characterization of metabotropic glutamate receptors in several types of brain cells in primary cultures.

Several cDNAs coding for metabotropic glutamate receptors (mGluR1-7) have now been isolated. mGluR1 and -5 are positively coupled to phospholipase C, whereas mGluR2, -3, -4, -6, and -7 are negatively coupled to adenylyl cyclase (AC) when they are expressed in Chinese hamster ovary or baby hamster kidney cells. However, the exact transduction mechanisms of these receptors in their natural environment remain to be determined.

Regional distribution and ontogeny of 5-HT4 binding sites in rodent brain.

We have investigated the regional distribution of 5-hydroxytryptamine4 (5-HT4) receptor binding sites in the adult guinea pig, rat and mouse brain using the specific 5-HT4 antagonist [3H]GR113808 as a radioligand. The developmental changes in the expression of these binding sites were also investigated quantitatively in the rat brain (gestational days 16 and 19; postnatal days 1, 3, 7, 9, 12 and 21). In order to compare previously obtained data on primary cultures, semi-quantitative analysis was also performed during mouse brain ontogeny (postnatal days 1, 7 and 11).