Research the Rainnie lab, based at the Yerkes National Primate Research Center, has been driven by my long-time interest in understanding the cellular processes that contribute to the perception of emotion. Emotion is often regarded as a psychological response rather than a physiological process, and yet, it exists simply because of an interaction between complex molecular, biochemical, and biophysical processes. Consequently, research in the Rainnie lab has focused on an investigation of the cellular and neurophysiological mechanisms underlying emotional aspects of cognition, with emphasis on the role of the extended amygdala in fear conditioning and extinction as well as anxiety-like behavior.
Multiple techniques ranging from molecular biology, through in vitro whole-cell patch clamp recording from visually identified neurons, to in vivo multiunit recording from freely moving rats are employed to examine the functional and neurochemical connectivity of the extended amygdala in an attempt to create a functional map of the intrinsic circuitry, and to determine how sensory information gains affective weight within this structure. By understanding how sensory information is processed in the extended amygdala, our ultimate objective is to shed light on the cellular processes that may contribute to the development of mood disorders such as depression, generalized anxiety disorder, panic disorder, and post traumatic stress disorder.
The combination of molecular genetic techniques and whole-cell patch clamp slice physiology offers a unique opportunity to examine the genetic phenotypes of distinct neuronal populations within the BLA. The goal of these studies is to use targeted manipulations of gene products within specific cell types to allow us to understand how individual genes within these cell types act to influence the normal function of the BLA, and ultimately their influence on affective behavior.
We are accomplishing these goals, primarily through the use of lentiviral vector-mediated gene transfer, and single cell reverse transcriptase PCR (sRT-PCR). With these techniques we can: 1) use lentiviral transfections of cell-specific promoters driving the expression of GFP to identify and record the physiological properties of distinct subpopulations of neurons within the BLA; 2) use the same system to over-express, introduce new, or knock-down, specific genes within distinct cell populations; and 3) use sRT-PCR to examine the genetic phenotype of specific neurons from physiologically distinct cell types. We can then characterize their mRNA expression pattern for individual neurotransmitters and their cognate receptors, second messenger systems, as well as ion channels and their chaperone proteins. This data can then be used to inform subsequent gene transfer experiments.
Specific projects include:
Promoter-based functional mapping of amygdala microcircuitry
Using dominant-negative gene constructs to examine the role of calcium-dependent potassium SK channels in the regulation of neural activity in the basolateral amygdala (BLA), and it's relation to emotion, cognition, and schizophrenia
Single cell RT-PCR analysis of neurotransmitter receptor subtypes in the BLA and BNST
Genetically targeted optical control of neural activity
Immunohistochemical, cytochemical, and tract tracing techniques are being used in combination with single cell morphological reconstructions to map the intrinsic and extrinsic connections of the basolateral amygdala (BLA) and the bed nucleus of the stria terminalis (BNST).
The focus of this research is to develop functional anatomical maps of the BLA and BNST in order to get a better understanding of their structure-function relationships. In particular, we hope to identify distinct cellular phenotypes within each region, examine the nature and extent of the connections between these cell types, and finally determine the level of interaction between these cell types and extrinsic afferent input, such as that arising from the brainstem monoamine systems.
Specific projects include:
An examination of the relative distribution of serotonin receptor subtypes in physiologically identified interneurons of the basolateral amygdala (BLA)
An examination of the morphological properties of three physiologically distinct subpopulations of neuron in the bed nucleus of the stria terminalis (BNST)
Mapping intrinsic neural connections in the BLA of rat and primate
Evaluating the relative expression of serotonin (5HT) receptor subtypes in define cell populations of the BNST using dual fluorescent in situ hybridization (FISH).
Evaluating the effects of chronic restraint stress on the expression of 5HT receptor subtypes in defined cell populations of the BNST using dual fluorescent in situ hybridization (FISH).
These studies use in vitro whole-cell patch clamp recording techniques to examine the physiological properties of individual neurons, as well as the local network properties of groups of neurons in the BLA and BNST. Viable sections of the rat brain, including the BLA and BNST, can be maintained for several hours in an oxygenated artificial cerebrospinal fluid. Individual neurons are visualized within the slice using infra-red microscopy and targeted for recording. Current- and voltage-clamp techniques are used to identify electrophysiologically distinct cell types in each region, and subsequently to determine their expression patterns for ion channels and neurotransmitter receptors.
Within the slice, the BLA and BNST retain much of their intrinsic and, to a lesser extent, extrinsic connections. Hence, we also examine the neurotransmitters involved in local synaptic connectivity by stimulating pairs of adjacent neurons, or in extrinsic synaptic connectivity by stimulating the afferent pathways to these neurons. The focus of these studies is to investigate the effects of local release of monoamine neurotransmitters and stress hormones on the physiological properties of BLA and BNST neurons
Specific projects include:
An examination of the cellular mechanisms contributing to dopaminergic enhancement of fear conditioning in the rat basolateral amygdala (BLA).
An examination of the cellular and network properties affecting serotonergic modulation of network excitability in the BLA.
An examination of the role of intrinsic membrane currents and synaptic potentials in the synchronization of BLA projection neurons.
An examination of the physiological properties of neurons in the primate BLA.
Identification of physiologically disctinct neuronal subtypes in the anterolateral bed nucleus of the stria terminalis (BNST).
An examination of the consequences of cross-talk between corticotropin releasing factor (CRF) receptors and serotonin receptors in the BNST
An examination of muscarinic receptor modulation of neural activity and synaptic transmission in the BNST.
A recent addition to the Rainnie laboratory is our in vivo multi-electrode recording set-up, which is showing great promise as a tool to bring systems-level understanding and behavioral relevance to the findings from our slice electrophysiology studies. By recording simultaneously in the BLA and infralimbic cortex (IL) of freely moving rats, we hope to demonstrate the role of coherent oscillations between these two structures in affect and emotional learning. Of particular interest is our collaboration with Dr. Helen Mayberg in which we hope to elucidate the neuronal mechanisms underlying the efficacy of deep brain stimulation (DBS) as a novel clinical therapy for treatment-resistant depression.
Specific projects include:
Characterization of neural activity in the BLA and IL of freely moving rats by recording single unit activity and local field potentials (LFPs).
Evaluating the role of coherence and synchronized neural oscillations in affective binding.
Monitoring changes in neural activity during fear conditioning and extinction training.
Attempting to modify those processes using microstimulation distributed across the multi-electrode recording array.
Modeling deep brain stimulation (DBS) for treatment resistant depression in order to identify its mechanisms.
Designing custom software to regulate DBS stimulation in unique ways.