Anandamide was the first of five endocannabinoids identified. It is an ethanolamide of arachidonic acid and it is an endogenous ligand for the CB1 receptor (CB1R) in the brain. Other endocannabinoids are 2-arachidonoyl glycerol (2-AG), noladin ether, virhodamine, and N-arachidonoyl dopamine. Dopamine is neurotransmitter and the precursor for two other neurotransmitters: norepinephrine and epinephrine. Dopamine is biosynthesized in two enzymatic steps from the amino acid tyrosine with an intermediate metabolite L-DOPA. Dopamine functions in many important brain circuits. Serotonin, which is also known as 5-hydroxytryptamine (5-HT), is a monoamine neurotransmitter synthesized from tryptophan in two enzymatic steps. Serotonin functions in the central nervous system (CNS) and gut. In the CNS, serotonin modulates a range of behaviors. In the gut, serotonin controls smooth muscle cell contraction. The cannabinoid receptor 1 is a G protein-coupled receptor that couples to the G i/o family. This receptor is abundant in the mammalian brain. The receptor is commonly associated with drug abuse mechanisms and is the known target for many agonists and antagonists among them marijuana. The natural agonists are termed endocannabinoids and can be released from the presynapse as a neurotransmitter. The cannabinoid receptor 1 also is important in development where it is expressed in neural progenitors. The dopamine 2 receptor (D2R) is a heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptor (GPCR) that leads to activation of Gi/o proteins, which inhibit adenylyl cyclase. D2R is found in nerve terminals and secretory cells. Drd2 are D2 dopamine receptors that couple to Gi/o and were implicated to induce neurite outgrowth in various in-vitro experiments. Neurons obtained from embryonic rats were stimulated with dopamine, and it was shown that the dopamine D2 receptors, are responsible for dendrite and axon elongation. 5-hydroxytryptamine receptors type 1B are G protein-coupled receptors that couple to Gi/o family of G proteins. Desensitization of the receptor may be mediated by its phosphorylation. This receptor is abundant in the fetal liver, dorsal root ganglia (DRG) neurons, and spinal cord. G beta/gamma The a, b, and g subunits of the heterotrimeric guanine nucleotide-binding protein (G protein) form a trimer and are associated in their inactive state. Activation of the G protein receptor (GPCR) results in a conformational change of the receptor that causes exchange of GDP for GTP by the a subunit, and the dissociation of the bg subunits from the a subunit. The bg subunits can then activate downstream signaling pathways. Gai is the a subunit of the heterotrimeric guanine nucleotide-binding protein inhibitory (Gi) complex. It inhibits adenylyl cyclase (AC) and decreases cellular adenosine 3',5'-monophosphate (cAMP) levels. The Gao family is homologous to the Gaii family. Both Gao and Gai can often bind to the same G protein-coupled receptors (GPCRs) but have different effectors. The downstream effectors of Go are much less understood compared with the effectors of Gi and Gs, which inhibit or stimulate adenylyl cyclases, respectively. Go can directly bind to the GTPase activating protein Rap1GAP. Such binding promotes Rap1GAP degradation by the proteasomal pathway. Upon receptor stimulation, more Gao is released to interact with Rap1GAP, resulting in Rap1 activation as Rap1GAP negatively regulates Rap1 activity by promoting its GAP activity. The adaptor subunit of PI3K (Phosphoinositide 3-kinase class Ia) p85 can be located at the vicinity of activating natural killer (NK) receptors at least through the DNAX-activating protein of 10 kDa (DAP10) adaptor protein, which contains a YxxM motif involved in the recognition of the Src homology 2 (SH2) domains of p85 p110 phosphatidylinositol 3-kinase (PI3K). PI3K is activated downstream activating NK receptors using immunoreceptor tyrosine-based activation motif (ITAM)-bearing adaptor proteins or DAP10 protein. PI3K inhibitors can block cytotoxicity and cytokine production in NK cells, thus regulating NK cell functions. At least two signaling pathways, dependant on the state of Syk kinase activation, can activate PI3K activity in NK cells. Phospholipase C-b (PLC-b) is activated by the asubunit of the q type of heterotrimeric guanine nucleotide-binding protein (Gaq) and also by Gbg. PLC-b has multiple isoforms. PLC-b1 and 3 are stimulated by Gaq, and PLC-b2 is stimulated by Gbg subunits. PLCs are soluble proteins that are partly cytosolic and partly associated with membrane. Activated PLC hydrolyzes phosphatidylinositol bisphosphate (PIP2) to inositol trisphosphate (IP3) and diacylglycerol (DAG). Adenylyl cyclases (ACs) were the first identified effectors for heterotrimeric guanine nucleotide-binding proteins (G proteins). Activation of ACs results in elevated levels of intracellular adenosine 3',5'-monophosphate (cAMP). The AC5 isoform is inhibited by the Gai subunit. Downstream effectors of adenylyl cyclase are the serine/threonine protein kinase PKA and the Rap guanine nucleotide exchange factor (GEF) Epac. The AC5 isoform is used in this pathway because it is known to be inhibited by Gi in mammalian neurons. Rap1 GTPase-activating protein 1 (Rap1GAP) is a GAP for the Ras-related protein Rap1. Rap1GAP converts Rap1 to its inactive GDP-bound state by stimulating GTPase activity. Rap1GAP is localized in the Golgi and the interleaf membrane, and is highly expressed in the brain and kidney. Rap1gap is inhibited by G-alpha-o. The inhibition of Rap1gap by G-alpha-o is achieved through direct interaction with G-alpha-o that leads Rap1gap degradation by the proteosome. The exact isoform that was shown to be degraded was Rap1GAPII. Phosphatidylinositol-3,4,5-trisphosphate [PI(3,4,5)P3] is a lipid product of phosphoinositide 3-kinase (PI3K) produced by phosphorylation of phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] at the 3' position of the inositol ring. Diacylglycerol (DAG) is formed from the hydrolysis of phosphatidylinositol bisphosphate (PIP2). DAG activates protein kinase C (PKC) by binding to its regulatory domain. Adenosine 3',5'-monophosphate (cAMP) is synthesized from adenosine triphosphate (ATP) by the plasma membrane-bound enzyme adenylyl cyclase (AC), and it is rapidly destroyed by cAMP phosphodiesterases. Phosphoinositide-dependent kinase 1 (PDK1) localizes to the plasma membrane by binding to the lipid products of phosphoinositide 3-kinase (PI3K), phosphatidylinositol-3,4,5-trisphosphate (PI345P3). This enzyme can phosphorylate critical Thr residue in the activation loop of various protein kinases in the protein kinase C (PKC) and protein kinase B (also known as Akt) superfamilies. This phosphorylation is an essential step in activation of these enzymes. Integrin-linked kinase (ILK) is a serine-threonine kinase encoded by a gene on chromosome 7. Ilk is highly expressed in heart followed by skeletal muscle, pancreas, and kidney, and weakly expressed in placenta, lung and liver. Ilk associates with the cytoplasmic domain of beta integrins can be activated by phosphatidylinositol-(3,4,5)-trisphosphate (PIP3) and is a kinase that can activate Akt. RasGap1 also known as SynGAP is a GTPase activating protein with an SH3 binding domain encoded by a gene on chromosome 17. This protein functions in neuronal development and it is selectively expressed in brain and is highly enriched at excitatory synapses. RasGAP1 stimulates the GTPase activity of Ras and as such negatively regulates Ras activity. RasGAP1 interacts with the PDZ domains of PSD-95 and SAP102. RasGRP1 (Ras guanine nucleotide releasing protein) is a guanine nucleotide exchange factor (GEF) that is encoded by a gene on chromosome 2. RasGRP1 is expressed in the brain with higher expression in cerebellum, cerebral cortex and amygdala. It is also expressed in the hematopoietic system and in T cells. RasGRP1 is stimulated by diacylglycerol, a cleavage product of phoshatidylinositol (3,4,5)-triphosphate (PIP3). RasGRP1 stimulates Ras GTPase to contribute to activation of the mitogen-activated protein kinase (MAPK) pathway. Protein kinase A (PKA) is made up of regulatory (R) and catalytic (C) subunits. In the inactive state the complex exists in an R2C2 state. Upon adenosine 3',5'-monophosphate (cAMP) binding the R subunits dissociate from the two C subunits, which then phosphorylate downstream effectors on Ser-Thr residues. Typical targets include metabolic enzymes, channels, and transcriptional regulators. Crk is a proto-oncogene that is located on chromosome 11 and encodes an adaptor protein. Crk binds to several tyrosine-phosphorylated proteins through several SH2 and SH3 domains. Phosphatidylinositol-3,4,5-trisphosphate (PI345P3) binds to the N-terminal pleckstrin homology (PH) domain of Akt (also known as PKB). This interaction both localizes Akt to the plasma membrane in response to appropriate receptor activation and makes Thr308 available for phosphorylation by phosphoinositide-dependent kinase 1 (PDK1). Phosphorylation of Thr308 is an essential to the activation of the kinase activity of Akt. This is a small guanine nucleotide-binding protein with intrinsic GTPase activity. GDP-bound Ras is inactive and GTP-bound Ras is active. In this pathway, the small GTPase Ras is thought to be responsible for the initial activation of MAPK1,2. For neurite-outgrowth, sustained activation of MAPK1,2 is required. This may be mediated through Rap1. In this pathway, the neuronal Ras, N-Ras is used, but it is known that other isoforms of Ras are present in mammalian neurons and in undifferentiated neuronal cell lines. Rap1 is a small guanosine triphosphatase (GTPase) regulated by Ca2+ and adenosine 3',5'-monophosphate (cAMP). In this pathway, Rap1 is important for sustained activation of the MAPK pathway required for initiating neurite outgrowth. With 8 links, Rap1 is the most connected node in the pathway. Long duration of MAPK activation, a requirement for neurite-outgrowth, is largely mediated by Rap1 activation. Although there are inhibitory signals to Rap1 from the reduction of activity of the cAMP effectors Epac and PKA, other upstream signals, such as those from Src, and the reduction of Rap1GAP by increased degradation, contribute to the sustained activity of Rap1. c-Raf, is a serine-threonine MAPK kinase-kinase that is activated by Ras to transduce the signal to downstream MAPK1,2. The activity of c-Raf is balanced by positive signaling from Ras and negative signaling from the cAMP-PKA and PI3K-AKT pathways. PKA and AKT phosphorylate c-Raf to inhibit its catalytic activity. c-Raf is expressed in most tissues. B-Raf is a serine-threonine kinase that is the human homolog of the avian c-Rmil protooncogene encoding a 94-kD serine/threonine kinase detected in avian cells. B-Raf is binds two Zn2= ions per subunit. It is mostly present in the cytoplasm of brain and testis tissue. Ral guanine nucleotide dissociation stimulator is a Ral-guanine-nucleotide exchange factor (GEF) that converts inactive Ral GTPase to its active GTP-bound state. RalGDS directly interacts with activated Ras, linking the Ral and Ras pathways. RalGDS was shown to bind to the same domain of the Ras-Raf interaction domain, acting as a sequestering inhibitor for the activation of Raf by Ras. RalGDS is highly expressed in the brain cortex and the ovaries. Mitogen-activated protein kinase or extracellular signal-regulated kinase (MEK) is a protein kinase that is phosphorylated and activated by Raf. Once activated, MEK phosphorylates and activates mitogen-activated protein kinase (MAPK) 1 and MAPK2. MEK is a dual-specificity protein kinase that phosphorylates MAPK on Ser-Thr and on Tyr residues. In this pathway, the small GTPase Ral is activated by one of its GEFs Ralgds. Ral is capable of activating the protein tyrosine kinase Src when it is in its GTP bound active form. The Ral GTPase has two isoforms: RalA and RalB, both expressed in neuronal cell lines with overlapping functions. Mitogen activated protein kinase (MAPK) 1 and 2 are members of the MAPK signaling family. MAPK 1,2 are, respectively, 42- and 44-kDa phosphoproteins that are activated upon phophorylation by the upstream signaling molecule MAPK kinase (MEK). MAPK 1,2 are Ser-Thr kinases that are able to phosphorylate a large number of downstream molecules such as signaling components and transcription factors. Src is a tyrosine protein kinase that was shown to be activated upon cannabinoid stimulation of Neuro2A cells. Src phosphorylates the transcription factor Stat-3, an important transcriptional regulator of genes required to initiate the neurite outgrowth program. MSK1,also known as ribosomal protein S6 kinase alpha 5 (RPS6KA5), is a serine-threonine kinase that is expressed in high levels in the heart, brain and placenta, and less abundantly in lung, kidney, and liver. RPS6KA5 is known to bind Mg2+ and activate the transcription factor CREB in neuronal cells. STAT3 is a transcription factor that can be activated by phosphorylation. The activated protein dimerizes and translocates to the nucleus. The two protein kinases JNK and Src have been implicated in Stat3 activation. The Src phosphorylation targets tyrosine residues. Stat3 activation is required for turning on the gene regulatory program that initiates neurite outgrowth. C3G is a 130 to 140 kD guanine-nucleotide exchange factor (GEF) for Rap1 and R-Ras. C3G was originally identified by virtue of its binding to the SH3 domain of the adaptor protein Crk. Binding to Crk is mediated by a centrally located proline-rich region. C3G has also been reported to bind directly to Cas. Phosphorylation of C3G on tyrosine is reported to increase the nucleotide exchange activity for Rap1. This member of the basic-leucine zipper (bZIP) family of transcription factors binds the adenosine 3',5'-monophosphate (cAMP) response element (CRE), a sequence present in many viral and cellular promoters. Upon binding to CRE, CREB1 stimulates transcription by binding to DNA as a dimer. non-covalent interaction : association : protein/metabolite non-covalent interaction : association : protein/metabolite non-covalent interaction : association : protein/metabolite non-covalent interaction : dissociation non-covalent interaction : dissociation non-covalent interaction : dissociation non-covalent interaction : dissociation non-covalent interaction : dissociation non-covalent interaction : dissociation non-covalent interaction : dissociation non-covalent interaction : dissociation non-covalent interaction : dissociation non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein covalent modification : lipid phosphorylation covalent modification : hydrolysis covalent modification : cyclization non-covalent interaction : association : protein/protein : GTPase activation non-covalent interaction : association : protein/phospholipid non-covalent interaction : association : protein/phospholipid non-covalent interaction : association : protein/phospholipid non-covalent interaction : association : protein/phospholipid non-covalent interaction : association : protein/metabolite covalent modification : protein phosphorylation : serine/threonine-phosphorylation covalent modification : protein phosphorylation : serine/threonine-phosphorylation non-covalent interaction : association : protein/protein non-covalent interaction : nucleotide exchange : GTP for GDP non-covalent interaction : nucleotide exchange : GDP for GTP covalent modification : protein phosphorylation : serine/threonine-phosphorylation covalent modification : protein phosphorylation : serine/threonine-phosphorylation non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein covalent modification : protein phosphorylation : serine/threonine-phosphorylation non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein non-covalent interaction : association : protein/protein covalent modification : protein phosphorylation : serine/threonine-phosphorylation covalent modification : protein phosphorylation : serine/threonine-phosphorylation non-covalent interaction : nucleotide exchange : GDP for GTP covalent modification : protein phosphorylation : dual specificity protein phosphorylation non-covalent interaction : association : protein/protein covalent modification : protein phosphorylation : serine/threonine-phosphorylation covalent modification : protein phosphorylation : tyrosine protein phosphorylation covalent modification : protein phosphorylation : tyrosine protein phosphorylation covalent modification : protein dephosphorylation : serine/threonine-dephosphorylation non-covalent interaction : association : protein/protein