Abstracts

Rationale: Dysregulation of a key node in the hippocampal network, the dentate gyrus (DG), is central in medial temporal lobe epilepsy (MTLE), where decreased DG inhibition is sufficient for seizure onset and inhibition can terminate seizures. The supramammillary hypothalamus (SuM) has topographically extensive bilateral projections to DG granule cells (GCs), being the largest direct extrinsic inhibitory input to the DG. A subpopulation of neurons surprisingly co-releases GABA and glutamate (SuMglut/GABA) and provides dense direct input to GC proximal dendrites; other subpopulations control hippocampal theta activity that is known to correlate with a very low probability of hippocampal interictal discharges and seizures. The precise targets of all SuM cell types are unknown, the trajectory of fibers through the brain has not been described, the physiological role of SuMglut/GABA neurons is unclear, and the involvement of the SuM in MTLE is unresolved. Here we describe the anatomical basis of the SuM's control of theta activity and the dentate gyrus. Methods: Efferent connections of SuM^glut/GABA, SuM^glut/NOS and SuM^glut neurons were examined using cell-selective vector-based (AAV-FLEX-hrGFP) tracing in Vglut2-Cre, NOS1-Cre and Vgat-Cre mice, as well as trans-synaptic anterograde tracing from the SuM (AAV1-DIO-flp; AAVDJ-fDIO-YFP at target). Afferent connections were determined using cholera toxin beta subunit (CTb). Results: SuM^glut/GABA neurons course through the lateral hypothalamus without clear terminals innervating this region, run up through the diagonal band region into the fimbria and fornix to very densely innervate the proximal dendrites of DG GCs, with relatively lighter innervation of the hilar region and CA2. In contrast, glutamate-only and NOS1-containing SuM neurons innervate the medial and lateral basal forebrain regions (e.g. substantia innominata, septal area), as well as the cerebral cortex.  SuM glutamate neurons innervate septal and basal forebrain GABAergic neurons, but do not innervate GABAergic neurons in the hippocampus. SuM inputs predominantly arose from the subicular region, tegmental nuclei, parabrachial region and basal forebrain.  Conclusions: GABA/glutamate co-releasing SuM neurons principally target the hippocampus proper, explaining our previously described finding of little influence on sleep-wake or hippocampal theta activity. These neurons are anatomically positioned to exert potent control over dentate granule cells, a key hippocampal input region. In contrast, non-GABA containing groups innervated regions that are critical for hippocampal theta activity, particularly the lateral septal area, with reciprocal connection with regions involved in sleep-wake control, including GABAergic basal forebrain neurons, again explaining our prior physiological results. Further ongoing work examines corresponding physiology, the role of these cell groups in MTLE, with further disambiguation of inputs using cell-selective glycoprotein-depleted rabies retrograde tracing. These specific cell populations may be selective targets for future treatments for MTLE as well as for disorders of arousal and memory. Funding: Support for this work was provided by the AAN (CRTF), and the Woodruff Foundation's support of the Emory Department of Neurology.