1 4 Microcircuits of the Amygdala Luke R. Johnson and Joseph E. LeDoux The amygdala was fi rst recognized as a distinct brain region in the early nine- teenth century (Swanson and Petrovich, 1998 ; LeDoux, 2007 ). The word amygdala, derived from the Greek, was meant to denote an almond-shaped structure identifi ed deep in the medial temporal lobe rostral to the ventral reaches of the hippocampus. Like most brain regions, the amygdala is not a single mass; rather, it is composed of distinct subareas or nuclei, each with anatomical and functional subdivisions (Pitkanen et al., 1997 ). These subdivi- sions are extensively connected with each other and other brain areas. The almond-shaped region that gives the amygdala its name was actually only one of these nuclei, the basal nucleus, rather than the whole structure. Today, the amygdala is best known for its role in emotional functions, especially fear, but it also contributes to memory and attention (McGaugh et al., 1996 ; Phelps and LeDoux, 2005 ). Unique microcircuits within the subdivisions of the amygdala are beginning to be identifi ed (Samson et al., 2003; Johnson and LeDoux, 2004 ; Samson and Pare, 2006 ; Johnson et al., 2008 ; Johnson et al., 2009 ). These are the subject of this review. First, though, we will discuss some organizational issues. What Is the Amgydala? Traditionally, the amygdala was viewed as consisting of an evolutionarily primitive division associated with the olfactory system (the cortico-medial region) and an evolutionarily newer division associated with the neocortex (the basolateral region) (Swanson and Petrovich, 1998 ; LeDoux, 2007 ). The cortico-medial region includes the cortical, medial, and central nuclei, while the basolateral region consists of the lateral, basal, and accessory 137 14-Shepherd-Ch14.indd 137 5/21/2010 3:30:20 PM 138 Handbook of Brain Microcircuits basal nuclei. However, a recent proposal by Swanson argues that the amygdala is neither a structural nor a functional unit, and instead consists of regions that belong to other regions or systems of the brain. For example, in this scheme, the lateral and basal amygdala are viewed as nuclear extensions of the cortex (rather than amygdala regions related to the cortex), while the cen- tral and medial amygdala are said to be ventral extensions of the striatum. This scheme has merit, but the present review focuses on the organization and function of the nuclei and subnuclei that are traditionally said to be part of the amygdala since most of the functions of the amygdala are understood in these terms. For example, extensive evidence suggests that Pavlovian fear conditioning depends on the amygdala. However, only select regions of the amygdala are involved. Specifi cally, the amygdala centric circuits underlying classical fear conditioning are well characterized (LeDoux, 2000 ; LeDoux, 2007 ). In this associative learning paradigm a conditioned stimulus (CS), usually an audi- tory tone, comes to elicit behavioral and autonomic signs of fear after occur- ring in conjunction with an aversive unconditioned stimulus (US), typically mild footshock. The CS and US converge in the lateral amygdala (Romanski et al., 1993 ; LeDoux, 2003 ). Fear conditioning is dependent on nociceptive inputs from the spinothalamic tract that terminate in the amygdala. These inputs may enter the amygdala via the thalamus or via the cortex (Shi and Davis, 2001 ). Upon later exposure to the CS, fear responses are expressed via connections from the lateral (LA) to the central (CE) amygdala. The LA and CE would still be important in fear conditioning even if the overall concept of the amygdala were eliminated. Neurons of the Amygdala Different amygdala nuclei have unique neuron types. The medial areas of the amygdala, including the CE and medial amygdala (M), are comprised of GABAergic projection neurons. These neurons are generally like the medium- sized spiny neurons (MSSNs) of the striatum in the basal ganglia. In contrast, the laterally located nuclei of the basoloteral complex (lateral [LA], basal [B], accessory basal [AB]) and the cortical amygdala nuclei (COA) are comprised predominantly of glutamatergic projection neurons ( Fig. 14.1 ). These projection neurons are similar to cortical projection neurons in morphology and electro- physiology. Unlike the cortex, the amygdala is not laminated, with the excep- tion of the cortical nuclei. The other glutamatergic neuron nuclei (LA, B, AB) do not show an immediately obvious structural organization. Given that the CE receives inputs from the LA and B, one immediate feature of amygdala micro- circuits is connectivity between cortical-like and basal ganglia–like neurons. The amygdala is more complex than the simple dichotomy of glutamater- gic and GABAergic projection neurons. The LA and B nuclei have been the 14-Shepherd-Ch14.indd 138 5/21/2010 3:30:20 PM 14: Microcircuits of the Amygdala 139 CS Sensory cortex mPFC Sensory thalamus CS CS LA CEm CEc “+” CEI Fear LAd No Fear “–” CE LAvm LAvl D “+” M L + BI ITC B V Md M Bm Mv ABpc ABmc AB CA F IGURE 14–1. Functional microcircuit of the lateral and central amygdala, which regulates fear and its extinction. Sensory and nociceptive circuits reach the lateral amygdala (LA) where synaptic plasticity occurs, storing key aspects of classically conditioned fear memory. For this memory to be behaviorally expressed, the amygdala output nucleus (medial division of central amygdala) CEm is activated indirectly via the GABAergic intercalated neurons (ITC). Aspects of the stored memory are transmitted from the LA to the basal (B) nucleus. In the B nucleus, neurons regulate the switch between the behavioral expression of fear or of its extinction. Behavioral expression is again routed through the CEm such that the original fear memory can remain intact but is overridden via the B to CEm circuit. GABAergic ITC play a key role in the gating amygdala microcircuits which express fear and fear extinction. most extensively studied (McDonald and Culberson, 1981 ; Rainnie et al., 1993 ; Pape et al., 2001 ; Sosulina et al., 2006 ; Mascagni et al., 2009 ). Earlier work reported up to seven separate populations of LA neurons (Faulkner and Brown, 1999 ). These potentially different populations of LA neurons were based on fi ring characteristics and morphology. More commonly reported are three unique populations: glutamatergic pyramidal and stellate projection neurons; and GABAergic local interneurons (Rainnie et al., 1991a ; Rainnie et al., 1991b ). Some data have suggested that only one population of glutamatergic principal neurons can be identifi ed in the LA. Pyramidal 14-Shepherd-Ch14.indd 139 5/21/2010 3:30:20 PM 140 Handbook of Brain Microcircuits neurons may look stellate-like when viewed from their ventral surface (Faber et al., 2001 ). With the exception of the cortical-amygdala nuclei, the amygdala has an apparent lack of neuron orientation and layering. However, important organization may be present in selective axonal connectivity. Within the amygdala as a whole there are at least three populations of GABAergic neurons. These are (1) GABAergic local interneurons throughout the amygdala. These interneurons are located within both the medial GABAergic nuclei and the lateral glutamatergic nuclei (Fig. 14.1). (2) The intercalated neurons (ITC), a population of small soma neurons positioned both medially and laterally (in the rat) to the LA that also use GABA as a neu- rotransmitter and topographically are part of the LA (the role of these neuron in an amygdala functional microcircuit is discussed in the section “The Intercalated Gate.” (3) The GABAergic projection neurons of the CE and M. These neurons are believed to be GABAergic medium sized spiny projection neurons (MSN) of same kind that comprise sections of the basal ganglia espe- cially the caudate and putamen (or striatum) (McDonald, 1982 ; Swanson and Petrovich, 1998 ). These neurons are different from the GABAergic local cir- cuit neurons. Glutamatergic principal and GABA interneurons of the LA and B differ in their spontaneous and maximal fi ring rates (Rainnie et al., 1991a ; Rainnie et al., 1991b ; Pare and Gaudreau, 1996 ; Royer and Pare, 2003 ; Sosulina et al., 2006 ). Principal neurons tend to be quiescent in vivo and in vitro unless activated by thalamic or cortical input or current injection. Following synap- tic activation, LA principal neurons can fi re action potentials up to 20 Hz. In contrast, presumed GABAergic interneurons are often spontaneously fi ring both in vivo and in vitro and can be induced to fi re up to 100 Hz. ITC neurons can fi re up to 30 Hz (Rainnie et al., 1991a ; Rainnie et al., 1991b ; Pare and Gaudreau, 1996 ; Sosulina et al., 2006 ; Woodruff and Sah, 2007 ). Like the cortex and hippocampus, the LA and B nuclei contain unique populations of local circuit neurons. The LA and B are known to contain excit- atory feedforward and feedback circuits (Samson et al., 2003 ; Johnson and LeDoux, 2004 ; Johnson et al., 2008 ). Increasing evidence indicates likely GABA circuits modulated by the known neuromodulators in a cell-specifi c manner (Rainnie, 1999 ; Pape, 2005 ; Rainnie et al., 2006 ; Mascagni and McDonald, 2007 ; Muller et al., 2007a ; Muller et al., 2007b ; Pinard et al., 2008 ). Moreover, these interneurons, especially the parvalbumin-positive GABA neurons, form inter- connected networks of inhibition and disinhibition (Muller et al., 2007a ; Muller et al., 2007b ; Woodruff and Sah, 2007 ; Sosulina et al., 2008 ). The Intercalated Gate Amygdala ITC neurons gate the fl ow of excitatory projections from the LA and BA to the CE. Discrete in vitro electrophysiological studies have identi- fi ed that LA and BA excitatory projection neurons make synaptic contact with 14-Shepherd-Ch14.indd 140 5/21/2010 3:30:20 PM 14: Microcircuits of the Amygdala 141 ITC GABA neurons, which in turn synapse onto GABAergic CE neurons (Pare and Smith, 1993 ; Pare and Smith, 1994 ; Pare et al., 2004 ). Importantly this ITC GABA input on CE neurons is able to shunt excitatory input from other exter- nal sources (Delaney and Sah, 2001 ; Sah and Westbrook, 2008 ).