Neurochemical Phenotypes Of The Afferent And Efferent Projections Of The Mouse Medial Habenula

The medial habenula (MHb) is a key bridge between limbic forebrain and midbrain monoaminergic centers. It is also conserved among different species. Although its exact behavioral function remains largely unknown, it is implicated in modulating as important behaviors as stress responses and circadian rhythm. Fundamental information such as the neurotransmitters in the afferent and efferent projections of the MHb remains unclear. (1)By combining retrograde tract tracing and genetic labeling of GABAergic neurons, we find that the medial septum (MS) and the nucleus of diagonal band (NDB) provideγ-aminobutyric acid (GABA)-ergic input to the MHb. (2)By anterograde tracing and immunostaining against a marker of glutamatergic synapses, we find that the projection from the triangular septal nucleus (TS) to the MHb has the capacity to release glutamate. This suggests that in addition to ATP, glutamate is another neurotransmitter for the TS to MHb projection. (3) By combining anterograde tracing and immunostaining, we find that the MHb neurons projecting to the interpeduncular nucleus (IPN) have the capacity to release glutamate. (4)Co-immunostaining indicates that vesicular transporters for glutamate and acetylcholine co-localize on the same axon terminals in the IPN; immuno-isolation of synaptic vesicles even shows that the two transporters are co-expressed in many synaptic vesicles. This suggests that, in addition to acetylcholine and substance P, glutamate is another neurotransmitter used by MHb projection neurons. MHb projection neurons may have the ability to co-release glutamate and acetylcholine. Our findings reveal the organization of key neurotransmitters for the MHb afferent and efferent projections, and lay framework for future functional studies of this pathway in the brain. Research on orphan G protein coupled receptors (oGPCRs) leads to the discovery of various new transmitters and benefits drug discovery. G protein coupled receptor 151(GPR151) is an orphan GPCR conserved in many different species, suggesting it is functional and binds to some endogenous ligand. We firstly took traditional "reverse pharmacology" and screeened dozens of molecules by calcium imaging. We next applied "orphan receptor strategy" to identify the ligand of GPR151 from animal brains and found peptide fractions which can specifically activate GPR151. By immuno-histochemistry, we found a "polarized" expression pattern of GPR151 in the axons of medial habenular projection neurons. The immunoreactivity co-localize with that of VAChT and VGLUT1, suggesting its role in cholinergic and glutamatergic innervations. To uncover the behavioral role of GPR151, we did behavioral analysis of the GPR151 knockout mice. We observed a stress-induced analgesia relating to GPR151. Our findings reveal that GPR151 may be retrogradely regulated by a peptidergic neuro-transmitter in the medial habenula-interpeduncular nucleus (MHb-IPN) axis and may serve its analgesic effect under stressful situations.