Abstract
Neurons producing serotonin (5-hydroxytryptamine, 5-HT) constitute one of the most widely distributed neuronal networks in the mammalian central nervous system (CNS) and exhibit a profuse innervation throughout the CNS already at early stages of development. Serotonergic neuron specification is controlled by a combination of secreted molecules and transcription factors such as Shh, Fgf4/8, Nkx2.2, Lmx1b and Pet1. In the mouse, Pet1 mRNA expression appears between 10 and 11 days post coitum (dpc) in serotonergic post-mitotic precursors and persists in serotonergic neurons up to adulthood, where it promotes the expression of genes defining the mature serotonergic phenotype such as tryptophan hydroxylase 2 (Tph2) and serotonin transporter (SERT). Hence, the generation of genetic tools based on Pet1 specific expression represents a valuable approach to study the development and function of the serotonergic system. Here, we report the generation of a Pet1210-Cre transgenic mouse line in which the Cre recombinase is expressed under the control of a 210 kb fragment from the Pet1 genetic locus to ensure a reliable and faithful control of somatic recombination in Pet1 cell lineage. Besides Cre-mediated recombination accurately occurred in the serotonergic system as expected and according to previous studies, Pet1210-Cre transgenic mouse line allowed us to identify novel, so far uncharacterized, Pet1 expression domains. Indeed, we showed that in the raphe Pet1 is expressed also in a non-serotonergic neuronal population intermingled with Tph2-expressing cells and mostly localized in the B8 and B9 nuclei. Moreover, we detected Cre-mediated recombination also in the developing pancreas and in the ureteric bud derivatives of the kidney, where it reflected a specific Pet1 expression. Thus, Pet1210-Cre transgenic mouse line faithfully drives Cre-mediated recombination in all Pet1 expression domains representing a valuable tool to genetically manipulate serotonergic and non-serotonergic Pet1 cell lineages.
Highlights
In mammals, neurons producing serotonin (5-hydroxytryptamine, 5-HT) are generated early during embryonic development in the ventral hindbrain and progressively cluster into B1–B9 raphe nuclei that project to the whole central nervous system (CNS), from the anterior brain to the spinal cord [1,2,3]
Southern Blot analysis on genomic DNA using a probe designed against the Kana/Neo DNA sequence allowed the identification of four independent Pet1210-Cre founders, three of which showed germline transmission, namely founder-female 3 (FF3), founder-female 9 (FF9) and founder-male 3 (FM3)
With immunohistochemistry data, few neurons expressing Pet1 but devoid of tryptophan hydroxylase 2 (Tph2) were detected in dorsal and caudal raphe B6 (Figure 3 d-d’’), B7 (Figure 3 c-c’’) and B1–B3 (Figure 3 e-e’’) nuclei whereas in B9 (Figure 3 a-a’’) and B8 (Figure 3 b-b’’) raphe nuclei a substantial number of non-serotonergic neurons resulted positive for Pet1 expression
Summary
Neurons producing serotonin (5-hydroxytryptamine, 5-HT) are generated early during embryonic development in the ventral hindbrain and progressively cluster into B1–B9 raphe nuclei that project to the whole central nervous system (CNS), from the anterior brain to the spinal cord [1,2,3]. Advances in mouse molecular genetics have brought new insights into the comprehension of the molecular cascade involved in serotonergic neuron specification [19,20,21,22,23,24], as well as of the projection network of raphe nuclei [11,25]. These studies have been crucial to reconsider serotonergic system, rather than a homogenous ensemble, a complex and heterogeneous population with distinct morphological, molecular and electrophysiological characteristics [22,24,25,26,27,28,29]
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