Synaptic Neurotransmission Depression in Ventral Tegmental Dopamine Neurons and Cannabinoid-Associated Addictive Learning

Drug addiction is an association of compulsive drug use with long-term associative learning/memory. Multiple forms of learning/memory are primarily subserved by activity- or experience-dependent synaptic long-term potentiation (LTP) and long-term depression (LTD). Recent studies suggest LTP expression in locally activated glutamate synapses onto dopamine neurons (local Glu-DA synapses) of the midbrain ventral tegmental area (VTA) following a single or chronic exposure to many drugs of abuse, whereas a single exposure to cannabinoid did not significantly affect synaptic plasticity at these synapses. It is unknown whether chronic exposure of cannabis (marijuana or cannabinoids), the most commonly used illicit drug worldwide, induce LTP or LTD at these synapses. More importantly, whether such alterations in VTA synaptic plasticity causatively contribute to drug addictive behavior has not previously been addressed. Here we show in rats that chronic cannabinoid exposure activates VTA cannabinoid CB1 receptors to induce transient neurotransmission depression at VTA local Glu-DA synapses through activation of NMDA receptors and subsequent endocytosis of AMPA receptor GluR2 subunits. A GluR2-derived peptide blocks cannabinoid-induced VTA synaptic depression and conditioned place preference, i.e., learning to associate drug exposure with environmental cues. These data not only provide the first evidence, to our knowledge, that NMDA receptor-dependent synaptic depression at VTA dopamine circuitry requires GluR2 endocytosis, but also suggest an essential contribution of such synaptic depression to cannabinoid-associated addictive learning, in addition to pointing to novel pharmacological strategies for the treatment of cannabis addiction

Synaptic Plasticity and NO-cGMP-PKG Signaling Regulate Pre- and Postsynaptic Alterations at Rat Lateral Amygdala Synapses Following Fear Conditioning

In vertebrate models of synaptic plasticity, signaling via the putative “retrograde messenger” nitric oxide (NO) has been hypothesized to serve as a critical link between functional and structural alterations at pre- and postsynaptic sites. In the present study, we show that auditory Pavlovian fear conditioning is associated with significant and long-lasting increases in the expression of the postsynaptically-localized protein GluR1 and the presynaptically-localized proteins synaptophysin and synapsin in the lateral amygdala (LA) within 24 hrs following training. Further, we show that rats given intra-LA infusion of either the NR2B-selective antagonist Ifenprodil, the NOS inhibitor 7-Ni, or the PKG inhibitor Rp-8-Br-PET-cGMPS exhibit significant decreases in training-induced expression of GluR1, synaptophysin, and synapsin immunoreactivity in the LA, while those rats infused with the PKG activator 8-Br-cGMP exhibit a significant increase in these proteins in the LA. In contrast, rats given intra-LA infusion of the NO scavenger c-PTIO exhibit a significant decrease in synapsin and synaptophysin expression in the LA, but no significant impairment in the expression of GluR1. Finally, we show that intra-LA infusions of the ROCK inhibitor Y-27632 or the CaMKII inhibitor KN-93 impair training-induced expression of GluR1, synapsin, and synaptophysin in the LA. These findings suggest that the NO-cGMP-PKG, Rho/ROCK, and CaMKII signaling pathways regulate fear memory consolidation, in part, by promoting both pre- and post-synaptic alterations at LA synapses. They further suggest that synaptic plasticity in the LA during auditory fear conditioning promotes alterations at presynaptic sites via NO-driven “retrograde signaling”

Localization of Mineralocorticoid Receptors at Mammalian Synapses

In the brain, membrane associated nongenomic steroid receptors can induce fast-acting responses to ion conductance and second messenger systems of neurons. Emerging data suggest that membrane associated glucocorticoid and mineralocorticoid receptors may directly regulate synaptic excitability during times of stress when adrenal hormones are elevated. As the key neuron signaling interface, the synapse is involved in learning and memory, including traumatic memories during times of stress. The lateral amygdala is a key site for synaptic plasticity underlying conditioned fear, which can both trigger and be coincident with the stress response. A large body of electrophysiological data shows rapid regulation of neuronal excitability by steroid hormone receptors. Despite the importance of these receptors, to date, only the glucocorticoid receptor has been anatomically localized to the membrane. We investigated the subcellular sites of mineralocorticoid receptors in the lateral amygdala of the Sprague-Dawley rat. Immunoblot analysis revealed the presence of mineralocorticoid receptors in the amygdala. Using electron microscopy, we found mineralocorticoid receptors expressed at both nuclear including: glutamatergic and GABAergic neurons and extra nuclear sites including: presynaptic terminals, neuronal dendrites, and dendritic spines. Importantly we also observed mineralocorticoid receptors at postsynaptic membrane densities of excitatory synapses. These data provide direct anatomical evidence supporting the concept that, at some synapses, synaptic transmission is regulated by mineralocorticoid receptors. Thus part of the stress signaling response in the brain is a direct modulation of the synapse itself by adrenal steroids

Afferents from the auditory thalamus synapse on inhibitory interneurons in the lateral nucleus of the amygdala.

Physiological studies suggest that afferents to the lateral nucleus of the amygdala (LA) from the auditory thalamus initiate feedforward inhibition [Li et al. (1996b)]. This model of neural processing requires that thalamic afferents synapse directly onto inhibitory interneurons. To determine whether such synaptic contacts occur, we combined anterograde tract tracing with interneuron immunocytochemistry. The anterograde tracer biotinylated dextran amine (BDA) was injected into the auditory thalamus. Inhibitory interneurons in the LA were identified using antibodies directed against gamma aminobutyric acid (GABA) or one of the calcium binding proteins (CBPs), parvalbumin (PARV), calbindin (CALB), or calretinin (CALR), since CBPs identify distinct populations of GABAergic cells within the amygdala. The distribution of GABAergic and CBP interneurons in each subregion of the LA was examined by light microscopy and the relationships between thalamo-amygdala terminals and interneurons were examined by confocal and electron microscopy. Immunoreactive cells were distributed in all three subdivisions of LA, except for CALR-ir neurons, which were sparse in the dorsal subregion and were found mainly in the ventromedial and ventrolateral subregions. Confocal microscopy revealed some thalamo-amygdala terminals in close proximity to LA interneurons, while electron microscopy showed that thalamo-amygdala terminals made direct synaptic contacts onto distal dendritic processes of inhibitory neurons. These data provide morphological evidence that thalamic afferents synapse directly onto inhibitory interneurons in LA, and are consistent with the possibility that inputs from the auditory thalamus initiate feedforward inhibition in LA. This architecture could play an important role in the suppression of background neural noise, thereby enhancing the response of LA cells to incoming auditory stimuli

Afferents from the auditory thalamus synapse on inhibitory interneurons in the lateral nucleus of the amygdala.

Physiological studies suggest that afferents to the lateral nucleus of the amygdala (LA) from the auditory thalamus initiate feedforward inhibition [Li et al. (1996b)]. This model of neural processing requires that thalamic afferents synapse directly onto inhibitory interneurons. To determine whether such synaptic contacts occur, we combined anterograde tract tracing with interneuron immunocytochemistry. The anterograde tracer biotinylated dextran amine (BDA) was injected into the auditory thalamus. Inhibitory interneurons in the LA were identified using antibodies directed against gamma aminobutyric acid (GABA) or one of the calcium binding proteins (CBPs), parvalbumin (PARV), calbindin (CALB), or calretinin (CALR), since CBPs identify distinct populations of GABAergic cells within the amygdala. The distribution of GABAergic and CBP interneurons in each subregion of the LA was examined by light microscopy and the relationships between thalamo-amygdala terminals and interneurons were examined by confocal and electron microscopy. Immunoreactive cells were distributed in all three subdivisions of LA, except for CALR-ir neurons, which were sparse in the dorsal subregion and were found mainly in the ventromedial and ventrolateral subregions. Confocal microscopy revealed some thalamo-amygdala terminals in close proximity to LA interneurons, while electron microscopy showed that thalamo-amygdala terminals made direct synaptic contacts onto distal dendritic processes of inhibitory neurons. These data provide morphological evidence that thalamic afferents synapse directly onto inhibitory interneurons in LA, and are consistent with the possibility that inputs from the auditory thalamus initiate feedforward inhibition in LA. This architecture could play an important role in the suppression of background neural noise, thereby enhancing the response of LA cells to incoming auditory stimuli

Calcium-permeable AMPA receptors mediate long-term potentiation in interneurons in the amygdala

Fear conditioning is a paradigm that has been used as a model for emotional learning in animals'. The cellular correlate of fear conditioning is thought to be associative N-methyl-D-aspartate (NMDA) receptor-dependent synaptic plasticity within the amygdala(1-3). Here we show that glutamatergic synaptic transmission to inhibitory interneurons in the basolateral amygdala is mediated solely by alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. In contrast to AMPA receptors at inputs to pyramidal neurons, these receptors have an inwardly rectifying current-voltage relationship, indicative of a high permeability to calcium(4 5), Tetanic stimulation of inputs to interneurons caused an immediate and sustained increase in the efficacy of these synapses. This potentiation required a rise in postsynaptic calcium, but was independent of NMDA receptor activation. The potentiation of excitatory inputs to interneurons was reflected as an increase in the amplitude of the GABAA-mediated inhibitory synaptic current in pyramidal neurons. These results demonstrate that excitatory synapses onto interneurons within a fear conditioning circuit show NMDA-receptor independent long-term potentiation. This plasticity might underlie the increased synchronization of activity between neurons in the basolateral amygdala after fear conditioning(6)