Axonal Varicosity Density as an Index of Local Neuronal Interactions

Diffuse transmission is an important non-synaptic communication mode in the cerebral neocortex, in which neurotransmitters released from en passant varicosities interact with surrounding cells. In a previous study we have shown that the cholinergic axonal segments which were in the microproximity with dopaminergic fibers possessed a greater density of en passant varicosities compared to more distant segments, suggesting an activity-dependent level of en passant varicosities in the axonal zone of interaction. To further evaluate this plastic relationship, the density of cholinergic varicosities was quantified on fiber segments within the microproximity of activated or non-activated pyramidal cells of the prefrontal cortex (mPFC). Repetitive 14 days patterned visual stimulation paired with an electrical stimulation of the cholinergic fibers projecting to the mPFC from the HDB was performed to induce persistent axonal plastic changes. The c-Fos early gene immunoreactivity was used as a neuronal activity marker of layer V pyramidal cells, labelled with anti-glutamate transporter EAAC1. Cholinergic fibers were labeled with anti-ChAT (choline acetyltransferase) immunostaining. The density of ChAT+ varicosities on and the length of fiber segments within the 3 µm microproximity of c-Fos positive/negative pyramidal cells were evaluated on confocal images. More than 50% of the pyramidal cells in the mPFC were c-Fos immunoreactive. Density of ChAT+ varicosities was significantly increased within 3 µm vicinity of activated pyramidal cells (0.50±0.01 per µm of ChAT+ fiber length) compared to non-activated cells in this group (0.34±0.001; p≤0.05) or control rats (0.32±0.02; p≤0.05). Different types of stimulation (visual, HDB or visual/HDB) induced similar increase of the density of ChAT+ varicosities within microproximity of activated pyramidal cells. This study demonstrated at the subcellular level an activity-dependent enrichment of ChAT+ varicosities in the axonal zone of interaction with other neuronal elements

Cholinergic Activation of M2 Receptors Leads to Context-Dependent Modulation of Feedforward Inhibition in the Visual Thalamus

The temporal dynamics of inhibition within a neural network is a crucial determinant of information processing. Here, the authors describe in the visual thalamus how neuromodulation governs the magnitude and time course of inhibition in an input-dependent way

The effective opening of nicotinic acetylcholine receptors with single agonist binding sites

We have identified a means by which agonist-evoked responses of nicotinic receptors can be conditionally eliminated. Modification of α7L119C mutants by the sulfhydryl reagent 2-aminoethyl methanethiosulfonate (MTSEA) reduces responses to acetylcholine (ACh) by more than 97%, whereas corresponding mutations in muscle-type receptors produce effects that depend on the specific subunits mutated and ACh concentration. We coexpressed α7L119C subunits with pseudo wild-type α7C116S subunits, as well as ACh-insensitive α7Y188F subunits with wild-type α7 subunits in Xenopus laevis oocytes using varying ratios of cRNA. When mutant α7 cRNA was coinjected at a 5:1 ratio with wild-type cRNA, net charge responses to 300 µM ACh were retained by α7L119C-containing mutants after MTSEA modification and by the ACh-insensitive Y188F-containing mutants, even though the expected number of ACh-sensitive wild-type binding sites would on average be fewer than two per receptor. Responses of muscle-type receptors with one MTSEA-sensitive subunit were reduced at low ACh concentrations, but much less of an effect was observed when ACh concentrations were high (1 mM), indicating that saturation of a single binding site with agonist can evoke strong activation of nicotinic ACh receptors. Single-channel patch clamp analysis revealed that the burst durations of fetal wild-type and α1β1γδL121C receptors were equivalent until the α1β1γδL121C mutants were exposed to MTSEA, after which the majority (81%) of bursts were brief (≤2 ms). The longest duration events of the receptors modified at only one binding site were similar to the long bursts of native receptors traditionally associated with the activation of receptors with two sites containing bound agonists

Dual Effect of Beta-Amyloid on α7 and α4β2 Nicotinic Receptors Controlling the Release of Glutamate, Aspartate and GABA in Rat Hippocampus

BACKGROUND: We previously showed that beta-amyloid (Aβ), a peptide considered as relevant to Alzheimer's Disease, is able to act as a neuromodulator affecting neurotransmitter release in absence of evident sign of neurotoxicity in two different rat brain areas. In this paper we focused on the hippocampus, a brain area which is sensitive to Alzheimer's Disease pathology, evaluating the effect of Aβ (at different concentrations) on the neurotransmitter release stimulated by the activation of pre-synaptic cholinergic nicotinic receptors (nAChRs, α4β2 and α7 subtypes). Particularly, we focused on some neurotransmitters that are usually involved in learning and memory: glutamate, aspartate and GABA. METHODOLOGY/FINDINGS: WE USED A DUAL APPROACH: in vivo experiments (microdialysis technique on freely moving rats) in parallel to in vitro experiments (isolated nerve endings derived from rat hippocampus). Both in vivo and in vitro the administration of nicotine stimulated an overflow of aspartate, glutamate and GABA. This effect was greatly inhibited by the highest concentrations of Aβ considered (10 µM in vivo and 100 nM in vitro). In vivo administration of 100 nM Aβ (the lowest concentration considered) potentiated the GABA overflow evoked by nicotine. All these effects were specific for Aβ and for nicotinic secretory stimuli. The in vitro administration of either choline or 5-Iodo-A-85380 dihydrochloride (α7 and α4β2 nAChRs selective agonists, respectively) elicited the hippocampal release of aspartate, glutamate, and GABA. High Aβ concentrations (100 nM) inhibited the overflow of all three neurotransmitters evoked by both choline and 5-Iodo-A-85380 dihydrochloride. On the contrary, low Aβ concentrations (1 nM and 100 pM) selectively acted on α7 subtypes potentiating the choline-induced release of both aspartate and glutamate, but not the one of GABA. CONCLUSIONS/SIGNIFICANCE: The results reinforce the concept that Aβ has relevant neuromodulatory effects, which may span from facilitation to inhibition of stimulated release depending upon the concentration used

Dopamine acting at D1-like, D2-like and α1-adrenergic receptors differentially modulates theta and gamma oscillatory activity in primary motor cortex

The loss of dopamine (DA) in Parkinson’s is accompanied by the emergence of exaggerated theta and beta frequency neuronal oscillatory activity in the primary motor cortex (M1) and basal ganglia. DA replacement therapy or deep brain stimulation reduces the power of these oscillations and this is coincident with an improvement in motor performance implying a causal relationship. Here we provide in vitro evidence for the differential modulation of theta and gamma activity in M1 by DA acting at receptors exhibiting conventional and non-conventional DA pharmacology. Recording local field potentials in deep layer V of rat M1, co-application of carbachol (CCh, 5 μM) and kainic acid (KA, 150 nM) elicited simultaneous oscillations at a frequency of 6.49 ± 0.18 Hz (theta, n = 84) and 34.97 ± 0.39 Hz (gamma, n = 84). Bath application of DA resulted in a decrease in gamma power with no change in theta power. However, application of either the D1-like receptor agonist SKF38393 or the D2-like agonist quinpirole increased the power of both theta and gamma suggesting that the DA-mediated inhibition of oscillatory power is by action at other sites other than classical DA receptors. Application of amphetamine, which promotes endogenous amine neurotransmitter release, or the adrenergic α1-selective agonist phenylephrine mimicked the action of DA and reduced gamma power, a result unaffected by prior co-application of D1 and D2 receptor antagonists SCH23390 and sulpiride. Finally, application of the α1-adrenergic receptor antagonist prazosin blocked the action of DA on gamma power suggestive of interaction between α1 and DA receptors. These results show that DA mediates complex actions acting at dopamine D1-like and D2-like receptors, α1 adrenergic receptors and possibly DA/α1 heteromultimeric receptors to differentially modulate theta and gamma activity in M1

MicroPET imaging of 5-HT1A receptors in rat brain: a test-retest [18F]MPPF study

International audiencePurposeEarlier studies have shown that positron emission tomography (PET) imaging with the radioligand [18F]MPPF allows for measuring the binding potential of serotonin 5-hydroxytryptamine1A (5-HT1A) receptors in different regions of animal and human brain, including that of 5-HT1A autoreceptors in the raphe nuclei. In the present study, we sought to determine if such data could be obtained in rat, with a microPET (R4, Concorde Microsystems). MethodsScans from isoflurane-anaesthetised rats (n = 18, including six test-retest) were co-registered with magnetic resonance imaging data, and binding potential, blood to plasma ratio and radiotracer efflux were estimated according to a simplified reference tissue model. ResultsValues of binding potential for hippocampus (1.2), entorhinal cortex (1.1), septum (1.1), medial prefrontal cortex (1.0), amygdala (0.8), raphe nuclei (0.6), paraventricular hypothalamic nucleus (0.5) and raphe obscurus (0.5) were comparable to those previously measured with PET in cats, non-human primates or humans. Test-retest variability was in the order of 10% in the larger brain regions (hippocampus, medial prefrontal and entorhinal cortex) and less than 20% in small nuclei such as the septum and the paraventricular hypothalamic, basolateral amygdaloid and raphe nuclei. ConclusionsMicroPET brain imaging of 5-HT1A receptors with [18F]MPPF thus represents a promising avenue for investigating 5-HT1A receptor function in rat

Anatomical and cellular localization of melatonin MT1 and MT2 receptors in the adult rat brain

The involvement of melatonin in mammalian brain pathophysiology has received growing interest, but information about the anatomical distribution of its two G-protein-coupled receptors, MT1 and MT2, remains elusive. In this study, using specific antibodies, we examined the precise distribution of both melatonin receptors immunoreactivity across the adult rat brain using light, confocal, and electron microscopy. Our results demonstrate a selective MT1 and MT2 localization on neuronal cell bodies and dendrites in numerous regions of the rat telencephalon, diencephalon, and mesencephalon. Confocal and ultrastructural examination confirmed the somatodendritic nature of MT1 and MT2 receptors, both being localized on neuronal membranes. Overall, striking differences were observed in the anatomical distribution pattern of MT1 and MT2 proteins, and the labeling often appeared complementary in regions displaying both receptors. Somadendrites labeled for MT1 were observed for instance in the retrosplenial cortex, the dentate gyrus of the hippocampus, the islands of Calleja, the medial habenula, the suprachiasmatic nucleus, the superior colliculus, the substantia nigra pars compacta, the dorsal raphe nucleus, and the pars tuberalis of the pituitary gland. Somadendrites endowed with MT2 receptors were mostly observed in the CA3 field of the hippocampus, the reticular thalamic nucleus, the supraoptic nucleus, the inferior colliculus, the substantia nigra pars reticulata, and the ventrolateral periaqueductal gray. Together, these data provide the first detailed neurocytological mapping of melatonin receptors in the adult rat brain, an essential prerequisite for a better understanding of melatonin distinct receptor function and neurophysiology