Neurotransmitter receptor localization: from autoradiography to imaging mass spectrometry.

Autoradiography is used to determine the anatomical distribution of biological molecules in human tissue and experimental animal models. This method is based on the analysis of the specific binding of radiolabeled compounds to locate neurotransmitter receptors or transporters in fresh frozen tissue slices. The anatomical resolution obtained by quantification of the radioligands has allowed the density of receptor proteins to be mapped over the last 40 years. The data yielded by autoradiography identify the receptors at their specific microscopic localization in the tissues and also in their native microenvironment, the intact cell membrane. Furthermore, in functional autoradiography, the effects of small molecules on the activity of G protein-coupled receptors are evaluated. More recently, autoradiography has been combined with membrane microarrays to improve the high-throughput screening of compounds. These technical advances have made autoradiography an essential analytical method for the progress of drug discovery. We include the future prospects and some preliminary results for imaging mass spectrometry (IMS) as a useful new method in pharmacodynamic and pharmacokinetic studies, complementing autoradiographic studies. IMS results could also be presented as density maps of molecules, proteins, and metabolites in tissue sections that can be identified, localized, and quantified, with the advantage of avoiding any labeling of marker molecules. The limitations and future developments of these techniques are discussed here.

Type-1 cannabinoid receptor activity during Alzheimer's disease progression.

The activity of CB1 cannabinoid receptors was studied in postmortem brain samples of Alzheimer's disease (AD) patients during clinical deterioration. CB1 activity was higher at earlier AD stages in limited hippocampal areas and internal layers of frontal cortex, but a decrease was observed at the advanced stages. The pattern of modification appears to indicate initial hyperactivity of the endocannabinoid system in brain areas that lack classical histopathological markers at earlier stages of AD, indicating an attempt to compensate for the initial synaptic impairment, which is then surpassed by disease progression. These results suggest that initial CB1 stimulation might have therapeutic relevance.

Distribution of lipids in human brain.

The enormous abundance of lipid molecules in the central nervous system (CNS) suggests that their role is not limited to be structural and energetic components of cells. Over the last decades, some lipids in the CNS have been identified as intracellular signalers, while others are known to act as neuromodulators of neurotransmission through binding to specific receptors. Neurotransmitters of lipidic nature, currently known as neurolipids, are synthesized during the metabolism of phospholipid precursors present in cell membranes. Therefore, the anatomical identification of each of the different lipid species in human CNS by imaging mass spectrometry (IMS), in association with other biochemical techniques with spatial resolution, can increase our knowledge on the precise metabolic routes that synthesize these neurolipids and their localization. The present study shows the lipid distribution obtained by MALDI-TOF IMS in human frontal cortex, hippocampus, and striatal area, together with functional autoradiography of cannabinoid and LPA receptors. The combined application of these methods to postmortem human brain samples may be envisioned as critical to further understand neurological diseases, in general, and particularly, the neurodegeneration that accompanies Alzheimer's disease.

Effects of 3,4-methylenedioxymethamphetamine ('Ecstasy') on the jaw-opening reflex and on the alpha-adrenoceptors which regulate this reflex in the anesthetized rat.

Bruxism, principally jaw clenching, is frequently observed in users of the recreational drug 3,4-methylenedioxymethamphetamine (MDMA). It has been suggested that during bruxism a reduction of the activity of oral protective reflexes occurs. In this study we investigated the effects of intravenously administered MDMA on the digastric electromyographic responses elicited by orofacial electrical stimulation in the rat. We also assessed the effects of either the administration of a single dose (20 mg kg(-1), s.c.) or repeated doses of MDMA (same dose, twice a day, for 4 d) on the jaw-opening reflex (JOR) and on the sensitivity of the alpha(2)-adrenoceptors which, in an inhibitory way, regulate it. Increasing doses of MDMA (1-29440 micro g kg(-1)) induced an incomplete inhibition of JOR and 50% inhibition (ED(50)) at 2550 micro g kg(-1); maximal inhibition was 88%. The repeated treatment with MDMA led to an enhancement of the inhibition of JOR induced by the alpha(2)-agonist, clonidine (ED(50) was reduced by 77%), indicating an increased sensitivity of the alpha(2)-adrenoceptors. This study shows that the intravenous administration of MDMA reduces the JOR while repeated doses of the drug enhance the inhibitory noradrenergic mechanisms which regulate the reflex. The results also allow speculation that a reduction of JOR may underlie the occurrence of episodes of bruxism in MDMA users.

Short-term effects of 3,4-methylenedioximethamphetamine on noradrenergic activity in locus coeruleus and hippocampus of the rat.

This study was undertaken to investigate the effects of the administration of 3,4-methylenedioxymethamphetamine (MDMA) on the locus coeruleus firing rate, on the sensitivity of the alpha(2)-adrenoceptors which regulate neuronal activity and on the in vivo tyrosine hydroxylase activity in hippocampus. The basal firing rate was not modified by either a single dose or repeated doses of MDMA, although the latter produced a shift to the right in the dose-response curve for clonidine-induced inhibition of the firing rate (ED(50) increased by 59%) and a reduction in tyrosine hydroxylase activity (20%) in the hippocampus. However, 8 days after the final dose alpha(2)-adrenoceptor sensitivity and tyrosine hydroxylase activity had returned to control values. Our results show a desensitization of alpha(2)-adrenoceptors in locus coeruleus and the existence of short-term changes in the noradrenergic system.