Specific Targeting of the Basolateral Amygdala to Projectionally Defined Pyramidal Neurons in Prelimbic and Infralimbic Cortex.

Adjacent prelimbic (PL) and infralimbic (IL) regions in the medial prefrontal cortex have distinct roles in emotional learning. A complete mechanistic understanding underlying this dichotomy remains unclear. Here we explored targeting of specific PL and IL neurons by the basolateral amygdala (BLA), a limbic structure pivotal in pain and fear processing. In mice, we used retrograde labeling, brain-slice recordings, and adenoviral optogenetics to dissect connectivity of ascending BLA input onto PL and IL neurons projecting to the periaqueductal gray (PAG) or the amygdala. We found differential targeting of BLA projections to PL and IL cortex. Activating BLA projections evoked excitatory and inhibitory responses in cortico-PAG (CP) neurons in layer 5 (L5) of both PL and IL cortex. However, all inhibitory responses were polysynaptic and monosynaptic BLA input was stronger to CP neurons in IL cortex. Conversely, the BLA preferentially targeted corticoamygdalar (CA) neurons in layer 2 (L2) of PL over IL cortex. We also reveal that BLA input is projection specific by showing preferential targeting of L5 CP neurons over neighboring L3/5 CA neurons in IL cortex. We conclude by showing that BLA input is laminar-specific by producing stronger excitatory responses CA neurons in L3/5 compared with L2 in IL cortex. Collectively, this study reveals differential targeting of the BLA to PL and IL cortex, which depends both on laminar location and projection target of cortical neurons. Overall, our findings should have important implications for understanding the processing of pain and fear input by the PL and IL cortex.

Highly differentiated cellular and circuit properties of infralimbic pyramidal neurons projecting to the periaqueductal gray and amygdala.

The infralimbic (IL) cortex is a key node in an inter-connected network involved in fear and emotion processing. The cellular and circuit-level mechanisms whereby IL neurons receive, filter, and modulate incoming signals they project onward to diverse downstream nodes in this complex network remain poorly understood. Using the mouse as our model, we applied anatomical labeling strategies, brain slice electrophysiology, and focal activation of caged glutamate via laser scanning photostimulation (glu-LSPS) for quantitative neurophysiological analysis of projectionally defined neurons in IL. Injection of retrograde tracers into the periaqueductal gray (PAG) and basolateral amygdala (BLA) was used to identify cortico-PAG (CP) and cortico-BLA (CA) neurons in IL. CP neurons were found exclusively in layer 5 (L5) of IL whereas CA neurons were detected throughout layer 2, 3, and 5 of IL. We also identified a small percentage of IL neurons that project to both the PAG and the BLA. We found that L5 CP neurons have a more extensive dendritic structure compared to L5 CA neurons. Neurophysiological recordings performed on retrogradely labeled neurons in acute brain slice showed that CP and CA neurons in IL could be broadly classified in two groups: neuronal resonators and non-resonators. Layer 2 CA neurons were the only class that was exclusively non-resonating. CP, CA, and CP/CA neurons in layers 3 and 5 of IL consisted of heterogeneous populations of resonators and non-resonators showing that projection target is not an exclusive predictor of intrinsic physiology. Circuit mapping using glu-LSPS revealed that the strength and organization of local excitatory and inhibitory inputs were stronger to CP compared to CA neurons in IL. Together, our results establish an organizational scheme linking cellular neurophysiology with microcircuit parameters of defined neuronal subclasses in IL that send descending commands to subcortical structures involved in fear behavior.

Probing NMDA receptor GluN2A and GluN2B subunit expression and distribution in cortical neurons.

The spatial distribution of N-methyl-d-aspartate receptor (NMDAR) subunits in layer 5 (L5) neurons of the medial prefrontal cortex (mPFC) is important for integrating input-output signals involved in cognitive functions and motor behavior. In this study, focal laser scanning photostimulation of caged glutamate, slice electrophysiology, and small peptide pharmacology, were used to map the distribution of functional GluN2A and GluN2B subunits of the NMDAR from L5 neurons of wild-type (WT) and GluN2A(-/-) mice. Focal uncaging of glutamate evoked spatially-restricted glutamatergic responses on various dendritic locations of pyramidal neurons in the mPFC. Analyses of the spatial arrangements of the GluN2A and GluN2B subunits were performed by comparing inhibition of glutamatergic responses in the presence of the GluN2A-selective pharmacological antagonist, NVP-AAM077 (NVP), and the GluN2B-selective peptidic antagonist, conantokin-G (con-G). We found that apical and basal expression and distribution of GluN2A and GluN2B were similar in L5 mPFC neurons of WT mice. However, the inhibition of glutamatergic responses by NVP in brain slices of GluN2A(-/-) mice were dramatically decreased, while con-G inhibition remained similar to that observed in WT brain slices. The data obtained show that expression and spatial arrangement of GluN2B subunits is independent of GluN2A in L5 neurons of the mPFC. These findings have important ramifications for NMDAR organization and function in L5 pyramidal neurons of the mPFC, and show that specific populations of NMDARs can be antagonized, while sparing other subgroups of NMDARs, thus preserving selective NMDAR functions, an important therapeutic advantage.