Neurite Branch Development of an Identified Serotonergic Neuron from Embryonic Helisoma: Evidence for Autoregulation by Serotonin

Abstract

Previous studies have shown that in select neurons, neurite outgrowth can be regulated by the same neurotransmitter that is synthesized and released by those neurons. However, it is not known whether such an autoregulatory mechanism is utilized during the normal course of nervous system development in either invertebrates or vertebrates. In the present study, we tested this hypothesis on the first pair of identified serotonergic neurons to be expressed in embryos of the pulmonate gastropod, Helisoma trivolvis. Embryonic neurons C1 (ENC1) elaborate a stereotyped pattern of neurite outgrowth prior to the differentiation of subsequent serotonergic neurons. Embryos were treated with either p-chlorophenylalanine (pCPA) or 5-hydroxytryptophan (5-HTP) to lower or raise embryonic serotonin content, respectively. High-performance liquid chromatography with electrochemical detection was used to measure the effects of these treatments on serotonin content, and serotonin immunohistochemistry was carried out to quantify the extent of neurite outgrowth of ENC1. Embryonic serotonin content was significantly reduced at both 24 and 48 hr after treatment with 0.02% pCPA, whereas dopamine levels were unchanged. Although the proximal neurite outgrowth of ENC1 appeared unaffected by the pCPA treatment at both of these time points, the distal outgrowth in the target cell region appeared more profuse. This effect on outgrowth was quantified by counting the number of neurite branch points, which was significantly increased both 24 and 48 hr after pCPA treatment. In contrast, 5-HTP treatment resulted in an increase in embryonic serotonin content and a significant decrease in the number of ENC1 branch points. Treatment with dopamine had no effect on the pattern of ENC1 neurite outgrowth. Together, these results support the hypothesis that a neuron may utilize its own transmitter in an autoregulatory fashion to regulate neurite formation during embryonic development.

Trajectories of brain development: point of vulnerability or window of opportunity?

“…For example, low concentrations, 5-methoxytryramine (0.1 -100 mM) inhibits serotonin uptake binding, whereas higher concentrations augment binding [191,228]. In culture, reduced serotonin increases dendritic branching at distal, but not proximal branch points, while augmented serotonin decreases neuronal branching [51].…”

Apical sensory neurones mediate developmental retardation induced by conspecific environmental stimuli in freshwater pulmonate snails

“…McCaman et al, 1984;Sakharov, 1991;Diefenbach et al, 1995;Fickbohm and Katz, 2000;Pires et al, 2000a), which is generally thought to be a result of neuronal activation. Methylated analogues and synthesis inhibitors are known to significantly reduce concentrations of respective monoamines as demonstrated by HPLC (e.g.…”

“…In contrast to marine invertebrates that mainly have free-swimming larvae, they develop inside transparent egg capsules and, hence, are well-suited for in vivo experimental studies of embryonic and larval development (Koss et al, 2003). Thus, in Helisoma trivolvis and Lymnaea stagnalis, various aspects of neuronal development and larval physiology have been studied (Goldberg and Kater, 1989;Goldberg et al, 1994;Diefenbach et al, 1991;Diefenbach et al, 1995;Croll and Voronezhskaya, 1996;Croll, 2000;Voronezhskaya and Elekes, 2003).…”

“…At the trochophore and veliger stages, only one pair of sensory neurones has been described (Diefenbach et al, 1998;Voronezhskaya et al, 1999). These neurones differ in their transmitter content: they express serotonin (5-HT) in Helisoma (Goldberg and Kater, 1989;Diefenbach et al, 1991;Diefenbach et al, 1995;Diefenbach et al, 1998;Voronezhskaya and Elekes, 1993;Koss et al, 2003), and dopamine (DA) and FMRFamide-related peptides in Lymnaea (Croll and Voronezhskaya, 1996;Voronezhskaya et al, 1999;Voronezhskaya and Elekes, 2003). Until the stage of late veliger these cells are the only monoaminergic neurones in both species.…”

“…First, endogenous monoamine level was augmented by incubation in its immediate biochemical precursor (Sakharov, 1991;Diefenbach et al, 1995). Conversely, the action of 5-HT and DA was reduced by treating the embryos with commonly used aromatic amino acid decarboxylase inhibitor 3-hydroxybenzylhydrazine (Treseder et al, 2003), specific inhibitor of tryptophan 5-hydroxylase activity L-pchlorophenylalanine (Baker et al, 1993;Diefenbach et al, 1995;Pani and Croll, 1998), methylated analogues of respective monoamines (Sloley and Orikasa, 1988) and receptor antagonists (Goldberg et al, 1994;Pavlova, 2001). Developmental changes were correlated with changes in anti-5-HT immunofluorescence in Helisoma, and glyoxylate-induced fluorescence of dopamine in Lymnaea within the first anterior neurones.…”