Cellular architecture and transmitter phenotypes of neurons of the mouse median raphe region

Abstract

The median raphe region (MRR, which consist of MR and paramedian raphe regions) plays a crucial role in regulating cortical as well as subcortical network activity and behavior, while its malfunctioning may lead to disorders, such as schizophrenia, major depression, or anxiety. Mouse MRR neurons are classically identified on the basis of their serotonin (5-HT), vesicular glutamate transporter type 3 (VGLUT3), and gamma-aminobutyric acid (GABA) contents; however, the exact cellular composition of MRR regarding transmitter phenotypes is still unknown. Using an unbiased stereological method, we found that in the MR, 8.5 % of the neurons were 5-HT, 26 % were VGLUT3, and 12.8 % were 5-HT and VGLUT3 positive; whereas 37.2 % of the neurons were GABAergic, and 14.4 % were triple negative. In the whole MRR, 2.1 % of the neurons were 5-HT, 7 % were VGLUT3, and 3.6 % were 5-HT and VGLUT3 positive; whereas 61 % of the neurons were GABAergic. Surprisingly, 25.4 % of the neurons were triple negative and were only positive for the neuronal marker NeuN. PET-1/ePET-Cre transgenic mouse lines are widely used to specifically manipulate only 5-HT containing neurons. Interestingly, however, using the ePET-Cre transgenic mice, we found that far more VGLUT3 positive cells expressed ePET than 5-HT positive cells, and about 38 % of the ePET cells contained only VGLUT3, while more than 30 % of 5-HT cells were ePET negative. These data should facilitate the reinterpretation of PET-1/ePET related data in the literature and the identification of the functional role of a putatively new type of triple-negative neuron in the MRR.

Connections of the laterodorsal tegmental nucleus with the habenular‐interpeduncular‐raphe system

“…Thus, DRL displays fairly specific connections, for example, with the IP and MnR, present findings (Lima et al, ), and has been selectively implicated in social stress (Crawford, Rahman, & Beck, ) and panic behavior (Johnson, Hollis, Moratalla, Lightman, & Lowry, ). Furthermore, our combined immunohistochemical/retrograde tracing findings suggest that all major neuronal populations present in the DR/MnR (Hioki et al, ; Sos et al, ) provide inputs to the LDTg, with a predominance of glutamatergic and GABAergic neurons, the later indicated indirectly by the elevated number of single labeled CTb neurons. An explicit role of the MnR in mechanisms of fear/anxiety has only been fully recognized during the last two decades.…”

“…For this purpose, injections of retrograde and anterograde tracers were made into the LDTg, LHb, IP, or MnR. Since LDTg (Wang & Morales, ), IP (Quina et al, ), and MnR (Sos et al, ) are now known to be neurochemically heterogeneous, we also characterized the transmitter phenotype of the projections between LDTg‐IP, and MnR‐LDTg. To this end, we combined retrograde tracing from IP or LDTg with immunofluorescence staining for choline acetyltransferase (ChAT), serotonin (5‐HT), and the Type 3 vesicular glutamate transporter (VGLUT3), and also with in situ hybridization for glutamate decarboxylase 67 kDA (GAD67), as well as the Type 2 vesicular glutamate transporter (VGLUT2).…”

Median raphe controls acquisition of negative experience in the mouse

“…Yet, it was not clear which neuronal pathway orchestrates the coordinated activation of these networks. Although the MRR was known to play a central role in processing negative experience (15,54), its known cell types did not project to the LHb, and the transmitter phenotypes and targets of almost 25% of the MRR neurons were not even known (22).…”

Regulatory Mechanisms Controlling Maturation of Serotonin Neuron Identity and Function

“…The number of Pet-1+ cells without detectable 5-HT identity was reported to be about 27% in the MRN while in the B7 and B6 portions of the DRN this number was only 1% (Pelosi et al, 2014). These findings were interpreted as evidence that not all Pet-1+ cells are 5-HT neurons (Sos et al, 2016). However, recent single cell RNA seq studies have revealed that all of the single Pet-1+ cells sequenced in the DRN and MRN express Tph2 , although some had a much lower number of Tph2 reads that other Pet-1+ cells (Okaty et al, 2015).…”