Dual medial prefrontal cortex and hippocampus projecting neurons in the paraventricular nucleus of the thalamus.

The paraventricular nucleus (PVT) of the midline thalamus is a critical higher-order cortico-thalamo-cortical integration site that plays a critical role in various behaviors including reward seeking, cue saliency, and emotional memory. Anatomical studies have shown that PVT projects to both medial prefrontal cortex (mPFC) and hippocampus (HC). However, dual mPFC-HC projecting neurons which could serve a role in synchronizing mPFC and HC activity during PVT-dependent behaviors, have not been explored. Here we used a dual retrograde adenoassociated virus (AAV) tracing approach to characterize the location and proportion of different projection populations that send collaterals to mPFC and/or ventral hippocampus (vHC) in rats. Additionally, we examined the distribution of calcium binding proteins calretinin (CR) and calbindin (CB) with respect to these projection populations in PVT. We found that PVT contains separate populations of cells that project to mPFC, vHC, and those that innervate both regions. Interestingly, dual mPFC-HC projecting cells expressed neither CR nor CB. Topographically, CB+ and CR+ containing cells clustered around dual projecting neurons in PVT. These results are consistent with the features of dual mPFC-vHC projecting cells in the nucleus reuniens (RE) and suggestive of a functional mPFC-PVT-vHC system that may support mPFC-vHC interactions in PVT-dependent behaviors.

Dual projecting cells linking thalamic and cortical communication routes between the medial prefrontal cortex and hippocampus.

The interactions between the medial prefrontal cortex (mPFC) and the hippocampus (HC) are critical for memory and decision making and have been specifically implicated in several neurological disorders including schizophrenia, epilepsy, frontotemporal dementia, and Alzheimer's disease. The ventral midline thalamus (vmThal), and lateral entorhinal cortex and perirhinal cortex (LEC/PER) constitute major communication pathways that facilitate mPFC-HC interactions in memory. Although vmThal and LEC/PER circuits have been delineated separately we sought to determine whether these two regions share cell-specific inputs that could influence both routes simultaneously. To do this we used a dual fluorescent retrograde tracing approach using cholera toxin subunit-B (CTB-488 and CTB-594) with injections targeting vmThal and the LEC/PER in rats. Retrograde cell body labeling was examined in key regions of interest within the mPFC-HC system including: (1) mPFC, specifically anterior cingulate cortex (ACC), dorsal and ventral prelimbic cortex (dPL, vPL), and infralimbic cortex (IL); (2) medial and lateral septum (MS, LS); (3) subiculum (Sub) along the dorsal-ventral and proximal-distal axes; and (4) LEC and medial entorhinal cortex (MEC). Results showed that dual vmThal-LEC/PER-projecting cell populations are found in MS, vSub, and the shallow layers II/III of LEC and MEC. We did not find any dual projecting cells in mPFC or in the cornu ammonis (CA) subfields of the HC. Thus, mPFC and HC activity is sent to vmThal and LEC/PER via non-overlapping projection cell populations. Importantly, the dual projecting cell populations in MS, vSub, and EC are in a unique position to simultaneously influence both cortical and thalamic mPFC-HC pathways critical to memory. SIGNIFICANCE STATEMENT: The interactions between mPFC and HC are critical for learning and memory, and dysfunction within this circuit is implicated in various neurodegenerative and psychiatric diseases. mPFC-HC interactions are mediated through multiple communication pathways including a thalamic hub through the vmThal and a cortical hub through lateral entorhinal cortex and perirhinal cortex. Our data highlight newly identified dual projecting cell populations in the septum, Sub, and EC of the rat brain. These dual projecting cells may have the ability to modify the information flow within the mPFC-HC circuit through synchronous activity, and thus offer new cell-specific circuit targets for basic and translational studies in memory.

Calretinin and calbindin architecture of the midline thalamus associated with prefrontal-hippocampal circuitry.

The midline thalamus bidirectionally connects the medial prefrontal cortex (mPFC) and hippocampus (HC) creating a unique cortico-thalamo-cortical circuit fundamental to memory and executive function. While the anatomical connectivity of midline thalamus has been thoroughly investigated, little is known about its cellular organization within each nucleus. Here we used immunohistological techniques to examine cellular distributions in the midline thalamus based on the calcium binding proteins parvalbumin (PV), calretinin (CR), and calbindin (CB). We also examined these calcium binding proteins in a population of reuniens cells known to project to both mPFC and HC using a dual fluorescence retrograde adenoassociated virus-based tracing approach. These dual reuniens mPFC-HC projecting cells, in particular, are thought to be important for synchronizing mPFC and HC activity. First, we confirmed the absence of PV+ neurons in the midline thalamus. Second, we found a common pattern of CR+ and CB+ cells throughout midline thalamus with CR+ cells running along the nearby third ventricle (3V) and penetrating the midline. CB+ cells were consistently more lateral and toward the middle of the dorsal-ventral extent of the midline thalamus. Notably, single-labeled CR+ and CB+ zones were partially overlapping and included dual-labeled CR+ /CB+ cells. Within RE, we also observed a CR and CB subzone specific diversity. Interestingly, dual mPFC-HC projecting neurons in RE expressed none of the calcium binding proteins examined, but were contained in nests of CR+ and CB+ cells. Overall, the midline thalamus was well organized into CR+ and CB+ rich zones distributed throughout the region, with dual mPFC-HC projecting cells in reuniens representing a unique cell population. These results provide a cytoarchitectural organization in the midline thalamus based on calcium binding protein expression, and set the stage for future cell-type specific interrogations of the functional role of these different cell populations in mPFC-HC interactions.