Parallel streams of raphe VGLUT3‐positive inputs target the dorsal and ventral hippocampus in each hemisphere

Abstract

The hippocampus (HP) receives neurochemically diverse inputs from the raphe nuclei, including glutamatergic axons characterized by the expression of the vesicular glutamate transporter type 3 (VGLUT3). These raphe‐HP VGLUT3 projections have been suggested to play a critical role in HP functions, yet a complete anatomical overview of raphe VGLUT3 projections to the forebrain, and in particular to the HP, is lacking. Using anterograde viral tracing, we describe largely nonoverlapping VGLUT3‐positive projections from the dorsal raphe (DR) and median raphe (MnR) to the forebrain, with the HP receiving inputs from the MnR. A limited subset of forebrain regions such as the amygdaloid complex, claustrum, and hypothalamus receives projections from both the DR and MnR that remain largely segregated. This highly complementary anatomical pattern suggests contrasting roles for DR and MnR VGLUT3 neurons. To further analyze the topography of VGLUT3 raphe projections to the HP, we used retrograde tracing and found that HP‐projecting VGLUT3‐positive neurons (VGLUT3HP) distribute over several raphe subregions (including the MnR, paramedian raphe, and B9 cell group) and lack co‐expression of serotonergic markers. Strikingly, double retrograde tracing experiments unraveled two parallel streams of VGLUT3‐positive projections targeting the dorsal and ventral poles of the HP. These results demonstrate highly organized and segregated VGLUT3‐positive projections to the HP, suggesting independent modulation of HP functions such as spatial memory and emotion‐related behavior.

Sex-specific modulation of the medial prefrontal cortex by glutamatergic median raphe neurons

“…The distinct effects of 5-HT antagonists suggest that the mechanisms underlying the increase in sIPSC frequency and amplitude are independent. Our experiments show that a small proportion of ChR2-expressing MRN (VGLUT3) neurons express Tph2, which is consistent with the recent reports on the presence of 5-HTergic markers in MRN (VGLUT3) neurons and the notable segregation of VGLUT3- and 5-HT–expressing neurons in the MRN ( 14 , 23 , 28 ). Our results showing MRN (VGLUT3) neuron–mediated glutamatergic activation of mPFC GABAergic neurons and the role of 5-HT in pyramidal neuron plasticity suggest a cooperative mechanism in MRN (VGLUT3) modulation of the mPFC.…”

“…Although our investigation of the effect of 5-HT is limited to its role in MRN (VGLUT3) neuron-mediated hyperpolarization and GABAergic plasticity in pyramidal neurons, these results strongly suggest the role of 5-HT release from MRN (VGLUT3) neurons on mPFC inhibition (10,14,19,28,(46)(47)(48). The hyperpolarization response in pyramidal neurons is mediated through 5-HT1A receptors, and these results are consistent with the previous studies (29,49).…”

“…Although MRN (VGLUT3) neuron–mediated glutamate release and the resulting activation of GABAergic neurons could affect mPFC GABAergic plasticity, the release of 5-HT from MRN (VGLUT3) neurons could also affect mPFC pyramidal neurons as a small proportion of the MRN (VGLUT3) neurons express a 5-HT marker, tryptophan hydroxylase 2 (Tph2; fig. S1) ( 14 , 28 ). Despite the lack of MRN (VGLUT3) neuron–mediated fast synaptic responses in pyramidal neurons, a 10-Hz light stimulation for 60 s produced a hyperpolarization response in 18 pyramidal neurons (6 neurons from 5 female mice and 12 neurons from 8 male mice) of 225 layer 5 pyramidal cells in MRN (VGLUT3)-ChR2 mice (127 neurons from 27 female + 98 neurons from 27 male mice) (Fig.…”

“…The hyperpolarization response in pyramidal neurons is mediated through 5-HT1A receptors, and these results are consistent with the previous studies ( 29 , 49 ). However, only a few pyramidal neurons show this hyperpolarization response, possibly due to the differential expression of 5-HT1A and excitatory 5-HT receptors such as 5-HT2A receptors or sparse MRN (VGLUT3) neuron–mediated 5-HT inputs in pyramidal neurons ( 28 , 49 ). Therefore, 5-HT1A receptors in the pyramidal neurons are unlikely to be responsible for MRN (VGLUT3) neuron–mediated GABAergic plasticity in female PL-mPFC.…”

“…A limitation of our study is the need for experiments to demonstrate the causal role of the PL-mPFC in MRN (VGLUT3)-mediated suppression of fear memory. Given the MRN (VGLUT3) inputs in the hippocampus and the medial septum, these brain areas also might play a role in the behavioral effects of MRN (VGLUT3) neuron activation ( 10 , 28 , 65 , 66 ). Therefore, future studies will be necessary to dissect the specific roles of these target brain areas in mediating the effect of MRN (VGLUT3) neurons on behavior.…”