Mechanisms underlying dual effects of serotonin during development of Helisoma trivolvis (Mollusca).

BACKGROUND: Serotonin (5-HT) is well known as widely distributed modulator of developmental processes in both vertebrates and invertebrates. It is also the earliest neurotransmitter to appear during neuronal development. In aquatic invertebrates, which have larvae in their life cycle, 5-HT is involved in regulation of stages transition including larval metamorphosis and settlement. However, molecular and cellular mechanisms underlying developmental transition in aquatic invertebrate species are yet poorly understood. Earlier we demonstrated that in larvae of freshwater molluscs and marine polychaetes, endogenous 5-HT released from the neurons of the apical sensory organ (ASO) in response to external stimuli retarded larval development at premetamorphic stages, and accelerated it at metamorphic stages. Here we used a freshwater snail Helisoma trivolvis to study molecular mechanisms underlying these dual developmental effects of 5-HT. RESULTS: Larval development of H. trivolvis includes transition from premetamorphic to metamorphic stages and shares the main features of metamorphosis with free-swimming aquatic larvae. Three types of 5-HT receptors (5-HT1-, 5-HT4- and 5-HT7-like) are functionally active at premetamorphic (trochophore, veliger) and metamorphic (veliconcha) stages, and expression patterns of these receptors and respective G proteins undergo coordinated changes during development. Stimulation of these receptors modulated cAMP-dependent regulation of cell divisions. Expression of 5-HT4- and 5-HT7-like receptors and their downstream Gs protein was down-regulated during the transition of pre- to metamorphic stage, while expression of 5-HT1 -like receptor and its downstream Gi protein was upregulated. In accordance with relative amount of these receptors, stimulation of 5-HTRs at premetamorphic stages induces developmental retardation, while their stimulation at metamorphic stages induces developmental acceleration. CONCLUSIONS: We present a novel molecular mechanism that underlies stage-specific changes in developmental tempo of H. trivolvis larvae in response to endogenous 5-HT produced by the neurons of the ASO. We suggest that consecutive changes in expression patterns of different receptors and their downstream partners in the course of larval development represent the molecular base of larval transition from premetamorphic (non-competent) to metamorphic (competent) state

5-HT1A receptor: its role in the regulation of different kinds of behavior

Brain serotonin (5-HT) is known to be involved in the control of a wide range of physiological functions as well as of different kinds of behavior. Such polyfunctionality of 5-HT is mediated by numerous 5-HT receptors. Currently, 14 different 5-HT receptor subtypes expressed in the mammals have been identified. The 5-HT1А receptor is one of the most extensively characterised members of the serotonin receptor family. Increased interest to the 5-HT1А receptor is based on (1) a key role in the autoregulation of the brain serotonergic system due to the postsynaptic and presynaptic localization, (2) a great body of data demonstrating implication of 5-HT1А receptor in the control of various physiological functions (3) involvement of 5-HT1А receptors in the mechanisms of depression, anxiety and suicide. The review describes literature and original data on factors affecting the expression and functional activity of 5-HT1А receptors and the involvement of 5-HT1А receptors in the regulation of normal and pathological behavior. The structure of the 5-HT1А receptor gene is described and new data on the posttranslational regulation of 5-HT1А receptor functional activity are provided. A special focus was given to the interaction between 5-HT1А and 5-HT7 receptors followed by heterodimer formation and the role of heterodimerization in the functional inactivation of the 5-HT1А receptor. The implication of 5-HT1А receptors in the regulation of aggressive behavior, catalepsy, anxiety, depression and hibernation was shown. Special attention is focused on the involvement of 5-HT1А receptors in the regulation of 1) fear-induced aggression towards man – the basis of domestication, 2) intermale aggression underling asocial behavior in men, 3) depression and in the mechanism of antidepressant action. The described data extend the idea on the 5-HT1А receptor as a key player in the brain 5-HT system

Expression of palmitoyl transferases in brain structures of mice genetically predisposed to depressive-like behavior

Most G-coupled receptors undergo posttranslational modifications. Among these modifications is S-palmitoylation, carried out by specialized enzymes palmitoyl transferases. Palmitoylation is the covalent attachment of a long-chain fatty acid, palmitate, to cysteine residues. It can influence receptor stability, transportation, and function. Obviously, malfunction of G-protein coupled receptors can cause various psychic disor­ders, including depression. However, no association between palmitoyl transferases and depressive-like behavior has been found hitherto. There is no informa­tion on brain structure specific features of palmitoyl transferase expression either. Here we investigate the expression of ZDHHC5, ZDHHC9, and ZDHHC21 palmi­toyl transferases in brain structures of ASC mice with genetic predisposition to depressive-like behavior in comparison with “nondepressive” CBA mice. Several brain region-specific features were detected in the immunodetection of palmitoyl transferase proteins. Western blot of the ZDHHC5 protein in the midbrain revealed two bands at 75 kDa and 55 kDa. Immuno­detection of ZDHHC21 palmitoyl transferase revealed two bands. One of them was visualized at 27 kDa in the frontal cortex and midbrain. The other, at 32 kDa in the hippocampus. Probing for ZDHHC9 also showed two bands in each of the midbrain and hippocampus, at 46 and 41 kDa. However, the expression of all investigated palmitoyl transferases in ASC mice with depressive-like behavior was almost identical to those in CBA mice. Thus, it was the first detection of brain region-specific features of the expression of investigated palmitoyl transferases. However, the study demonstrates that the genetic predisposition to depression-like behavior in ASC mice is not associated with changes in ZDHHC5, ZDHHC9, or ZDHHC21 palmitoyl transferase expression

Synaptic Remodeling Depends on Signaling between Serotonin Receptors and the Extracellular Matrix

Rewiring of synaptic circuitry pertinent to memory formation has been associated with morphological changes in dendritic spines and with extracellular matrix (ECM) remodeling. Here, we mechanistically link these processes by uncovering a signaling pathway involving the serotonin 5-HT7 receptor (5-HT7R), matrix metalloproteinase 9 (MMP-9), the hyaluronan receptor CD44, and the small GTPase Cdc42. We highlight a physical interaction between 5-HT7R and CD44 (identified as an MMP-9 substrate in neurons) and find that 5-HT7R stimulation increases local MMP-9 activity, triggering dendritic spine remodeling, synaptic pruning, and impairment of long-term potentiation (LTP). The underlying molecular machinery involves 5-HT7R-mediated activation of MMP-9, which leads to CD44 cleavage followed by Cdc42 activation. One important physiological consequence of this interaction includes an increase in neuronal outgrowth and elongation of dendritic spines, which might have a positive effect on complex neuronal processes (e.g., reversal learning and neuronal regeneration)

Spermidine protects from age-related synaptic alterations at hippocampal mossy fiber-CA3 synapses

Aging is associated with functional alterations of synapses thought to contribute to age-dependent memory impairment (AMI). While therapeutic avenues to protect from AMI are largely elusive, supplementation of spermidine, a polyamine normally declining with age, has been shown to restore defective proteostasis and to protect from AMI in Drosophila. Here we demonstrate that dietary spermidine protects from age-related synaptic alterations at hippocampal mossy fiber (MF)-CA3 synapses and prevents the aging-induced loss of neuronal mitochondria. Dietary spermidine rescued age-dependent decreases in synaptic vesicle density and largely restored defective presynaptic MF-CA3 long-term potentiation (LTP) at MF-CA3 synapses (MF-CA3) in aged animals. In contrast, spermidine failed to protect CA3-CA1 hippocampal synapses characterized by postsynaptic LTP from age-related changes in function and morphology. Our data demonstrate that dietary spermidine attenuates age-associated deterioration of MF-CA3 synaptic transmission and plasticity. These findings provide a physiological and molecular basis for the future therapeutic usage of spermidine

mTORC1 activity is supported by spatial association with focal adhesions

The mammalian target of rapamycin complex 1 (mTORC1) integrates mitogenic and stress signals to control growth and metabolism. Activation of mTORC1 by amino acids and growth factors involves recruitment of the complex to the lysosomal membrane and is further supported by lysosome distribution to the cell periphery. Here, we show that translocation of lysosomes toward the cell periphery brings mTORC1 into proximity with focal adhesions (FAs). We demonstrate that FAs constitute discrete plasma membrane hubs mediating growth factor signaling and amino acid input into the cell. FAs, as well as the translocation of lysosome-bound mTORC1 to their vicinity, contribute to both peripheral and intracellular mTORC1 activity. Conversely, lysosomal distribution to the cell periphery is dispensable for the activation of mTORC1 constitutively targeted to FAs. This study advances our understanding of spatial mTORC1 regulation by demonstrating that the localization of mTORC1 to FAs is both necessary and sufficient for its activation by growth-promoting stimuli

Serotonin 5-HT4 receptor boosts functional maturation of dendritic spines via RhoA-dependent control of F-actin

Activity-dependent remodeling of excitatory connections underpins memory formation in the brain. Serotonin receptors are known to contribute to such remodeling, yet the underlying molecular machinery remains poorly understood. Here, we employ high-resolution time-lapse FRET imaging in neuroblastoma cells and neuronal dendrites to establish that activation of serotonin receptor 5-HT4 (5-HT4R) rapidly triggers spatially-restricted RhoA activity and G13-mediated phosphorylation of cofilin, thus locally boosting the filamentous actin fraction. In neuroblastoma cells, this leads to cell rounding and neurite retraction. In hippocampal neurons in situ, 5-HT4R-mediated RhoA activation triggers maturation of dendritic spines. This is paralleled by RhoA-dependent, transient alterations in cell excitability, as reflected by increased spontaneous synaptic activity, apparent shunting of evoked synaptic responses, and enhanced long-term potentiation of excitatory transmission. The 5-HT4R/G13/RhoA signaling thus emerges as a previously unrecognized molecular pathway underpinning use-dependent functional remodeling of excitatory synaptic connections

Specific oligomerization of the 5-HT1A receptor in the plasma membrane

In the present study we analyze the oligomerization of the 5-HT1A receptor within living cells at the sub-cellular level. Using a 2-excitation Förster Resonance Energy Transfer (FRET) method combined with spectral microscopy we are able to estimate the efficiency of energy transfer based on donor quenching as well as acceptor sensitization between CFP-and YFP-tagged 5-HT1A receptors at the plasma membrane. Through the analysis of the level of apparent FRET efficiency over the various relative amounts of donor and acceptor, as well as over a range of total surface expressions of the receptor, we verify the specific interaction of these receptors. Furthermore we study the role of acylation in this interaction through measurements of a palmitoylation-deficient 5-HT1A receptor mutant. Palmitoylation increases the tendency of a receptor to localize in lipid rich microdomains of the plasma membrane. This increases the effective surface density of the receptor and provides for a higher level of stochastic interaction

The Transcription Factor Cux1 Regulates Dendritic Morphology of Cortical Pyramidal Neurons

In the murine cerebral cortex, mammalian homologues of the Cux family transcription factors, Cux1 and Cux2, have been identified as restricted molecular markers for the upper layer (II-IV) pyramidal neurons. However, their functions in cortical development are largely unknown. Here we report that increasing the intracellular level of Cux1, but not Cux2, reduced the dendritic complexity of cultured cortical pyramidal neurons. Consistently, reducing the expression of Cux1 promoted the dendritic arborization in these pyramidal neurons. This effect required the existence of the DNA-binding domains, hence the transcriptional passive repression activity of Cux1. Analysis of downstream signals suggested that Cux1 regulates dendrite development primarily through suppressing the expression of the cyclin-dependent kinase inhibitor p27Kip1, and RhoA may mediate the regulation of dendritic complexity by Cux1 and p27. Thus, Cux1 functions as a negative regulator of dendritic complexity for cortical pyramidal neurons