While I have found several good indications pointing to a rodent AOS-like organization of the lateral olfactory stream in larval Xenopus laevis, the similar ratio of MTCs innervating one or two glomeruli does not seem to fit to the hypothesis at first sight. Several of the Xenopus MTCs I observed, resemble MCs or TCs of the rodent MOB, innervating one glomerulus with their primary dendritic tuft (Nagayama et al., 2014; Pinching and Powell, 1971a). In rodents, this type of neuron has so far almost exclusively been described in the MOS in combination with cAMP-dependent signal transduction and OR-type receptor expression of their parental glomeruli (Munger et al., 2009; Nagayama et al., 2014). The lateral olfactory stream in Xenopus is devoid of glomeruli/ORN axons that use cAMP-dependent signal transduction (Gliem et al., 2013). It cannot be ruled out completely, that OR-type glomeruli with cAMP-independent signal transduction would be present in the LC. In fact, there are reports of an OR-expressing, non-ciliated ORN population, which projects to the AOB in rodents and uses TRPC2 mediated signal transduction (Lévai et al., 2006). It is imaginable that also in those ORNs, the OR would dictate uni-glomerular axonal wiring. Too few is known about this ORN species to draw any conclusions. Rodents are among the most commonly used animal models in olfactory research. However, their MOS and consequently, the MTC types are
109 mostly representative of air-borne odor detection systems (Bear et al., 2016; Munger et al., 2009). In zebrafish, most MTCs are innervating one glomerulus with their multiple primary dendrites (Fuller et al., 2006; Wanner et al., 2016). Consequently, zebrafish possess a cAMP-independent, amino acid-sensitive subsystem with uni-glomerular MTCs (Friedrich and Korsching, 1997; Fuller et al., 2006; Sato et al., 2005; Wanner et al., 2016). What could be the reason that some V2R-based olfactory subsystems exhibit mainly multi-glomerular MTCs (rodent AOB) whereas others are dominated by the uni-glomerular MTC type? A possible explanation might be that though they are structurally related, OlfC receptors and mammalian V2Rs are phylogenetically distant receptors (Alioto and Ngai, 2005, 2006). An intriguing hypothesis could be that the V2R receptor family's sequence diversity (including OlfC) results in different axonal wiring strategies depending on the individual receptor. ORNs that express OlfC receptors in zebrafish (and maybe other teleosts as well) could be an example of this clade's wiring logic in which axons coalesce to one glomerulus, and convey their input to a set of uni-glomerular MTCs. Later diverging ‘mammalian'-like V2Rs in rodents form multiple glomeruli enabling cross-channel integration from several glomeruli by multi-glomerular MTCs (Belluscio et al., 1999; Rodriguez et al., 1999; Takami and Graziadei, 1991; Wagner et al., 2006). It is tempting to hypothesize that the diversity in V2R repertoire in Xenopus containing both more ancestral fish-like and later diverging V2R variants (Hagino-Yamagishi et al., 2004; Syed et al., 2013) would result in co-existence of both wiring logics. In the principal cavity of larval Xenopus, more ancestral V2R variants are expressed (Syed et al., 2013). That would mean, this ancestral receptor subgroup could possibly be divided further by their uni- or multi-glomerular wiring strategy. It is imaginable that the previously discussed selective co-expression of V2Rs with a V2R group C member might be predictive for one of the wiring patterns. In larval Xenopus, a subset of V2Rs that are more similar to OlfC receptors could contribute to the uni-glomerular wiring logic, whereas other ancestral 'fish-like' but later diverging V2Rs might be the source of the multi-glomerular wiring logic. The number of glomeruli that were maximally innervated by MTCs in Xenopus in this study was four, and the innervation of two glomeruli per MTC was the most common variant (33%). This intermediate number of innervated glomeruli in amphibians compared to mammalian AMCs of the V2R domain (Larriva-Sahd, 2008; Takami and Graziadei, 1991) could lead to the impression, that there was a trend to increasing number of glomeruli, the later the V2R family diverged in vertebrates (Shi and Zhang, 2007). However, this linear relationship solely based on phylogeny seems very unlikely, since actually in the majority of other teleost fish investigated, MTCs form synapses with several glomeruli, often more than two (Dryer and Graziadei, 1994; Oka, 1983). Whether V2R-based axonal wiring is the basis of these MTC morphologies is not known. One way to interpret my results of the MTC morphological analysis is that at least two different wiring logics co-exist in the lateral olfactory system of larval Xenopus. One of them is more rodent AOS-like with MTC cross-channel integration from multiple glomeruli (Wagner et al., 2006). The other is based on singular glomerular
110 innervation from pre- and postsynapse in a more labeled line like configuration (Mombaerts, 2006; Mombaerts et al., 1996). Those different wiring patterns could be instructed by different groups of V2Rs that do not necessarily need to match the phylogenetic subgroups.
One could argue, that V1Rs might as well be suitable candidates for some of the observed wiring phenomena in Xenopus since they are supposedly using cAMP-independent signal transduction in rodents (Munger et al., 2009) and are found in the larval XenopusMOS (Syed et al., 2013). However, the variability of the glomerular map, as well as the high prevalence of multi-glomerular MTCs in the amino acid-sensitive mediolateral lobe of the LC, strongly suggests that at least a part of the Xenopus multi-glomerular MTC population might be receiving input from V2Rs. It cannot be excluded that V1R-based glomerular wiring patterns independently contribute to either the uni- or multi-glomerular MTC population. If Amphibian V1Rs were similar to mammalian V1Rs in that sense, the multi-glomerular phenotype of MTCs could occur preferentially (Wagner et al., 2006).
Schematic 20 ORN axonal wiring strategies and possible odorant receptors involved
111