In contrast to the single sensory surface present in teleost fishes, several spatially segregated subsystems with distinct molecular and functional characteristics define the mammalian olfactory system. However, the evolutionary steps of that transition remain unknown. Here we analyzed the olfactory system of an early diverging tetrapod, the amphibian Xenopus laevis, and report for the first time the existence of two odor-processing streams, sharply segregated in the main olfactory bulb and partially segregated in the olfactory epithelium of pre-metamorphic larvae. A lateral odor-processing stream is formed by microvillous receptor neurons and is characterized by amino acid responses and Gαo/Gαi as probable signal transducers, whereas a medial stream formed by ciliated receptor neurons is characterized by responses to alcohols, aldehydes, and ketones, and Gαolf/cAMP as probable signal transducers. To reveal candidates for the olfactory receptors underlying these two streams, the spatial distribution of 12 genes from four olfactory receptor gene families was determined. Several class II and some class I odorant receptors (ORs) mimic the spatial distribution observed for the medial stream, whereas a trace amine-associated receptor closely parallels the spatial pattern of the lateral odor-processing stream. Other olfactory receptors (some class I odorant receptors and vomeronasal type 1 receptors) and odor responses (to bile acids, amines) were not lateralized, the latter not even in the olfactory bulb, suggesting an incomplete segregation. Thus, the olfactory system of X. laevis exhibits an intermediate stage of segregation and as such appears well suited to investigate the molecular driving forces behind olfactory regionalization.

Extracellular purines and pyrimidines are important signaling molecules acting via purinergic cell-surface receptors in neurons, glia, and glia-like cells such as sustentacular supporting cells (SCs) of the olfactory epithelium (OE). Here, we thoroughly characterize ATP-induced responses in SCs of the OE using functional Ca2+ imaging. The initial ATP-induced increase of the intracellular Ca2+ concentration [Ca2+]i always occurred in the apical part of SCs and subsequently propagated toward the basal lamina, indicating the occurrence of purinergic receptors in the apical part of SCs. The mean propagation velocity of the Ca2+ signal within SCs was 17.10 +/- 1.02 microm/s. ATP evoked increases in [Ca2+]i in both the presence and absence of extracellular Ca2+. Depletion of the intracellular Ca2+ stores abolished the responses. This shows that the ATP-induced [Ca2+]i increases were in large part, if not entirely, due to the activation of G protein-coupled receptors followed by Ca2+ mobilization from intracellular stores, suggesting an involvement of P2Y receptors. The order of potency of the applied purinergic agonists was UTP > ATP > ATPgammaS (with all others being only weakly active or inactive). The ATP-induced [Ca2+]i increases could be reduced by the purinergic antagonists PPADS and RB2, but not by suramin. Our findings suggest that extracellular nucleotides in the OE activate SCs via P2Y2/P2Y4-like receptors and initiate a characteristic intraepithelial Ca2+ wave.

In the olfactory epithelium (OE) continuous neurogenesis is maintained throughout life. The OE is in direct contact with the external environment, and its cells are constantly exposed to pathogens and noxious substances. To maintain a functional sense of smell the OE has evolved the ability to permanently replenish olfactory receptor neurons and sustentacular cells lost during natural turnover. A cell population residing in the most basal part of the OE, the so-called basal cells (BCs), keep up this highly regulated genesis of new cells. The population of BCs is thought to include both the stem cells of the OE and various progenitor cells. In recent years a number of regulatory factors that positively and/or negatively regulate the proliferation within the OE have been identified, but a thorough comprehension of the complex interplay of these regulatory factors and the role of the different epithelial cell types is still illusive. Combining labeling techniques, immunohistochemistry, electron microscopy, functional calcium imaging, and a bromo-2 0 -deoxyuridine incorporation assay, we show for the first time that purinergic receptors are expressed in BCs of the OE of larval Xenopus laevis and that nucleotide-induced Ca 21 signaling in these cells is involved in the regulation of the cell turnover in the OE. Our data contribute to a better understanding of the regulation of the cell turnover in the OE in particular and also of how the proliferation of neuronal progenitor cells is regulated in general.