Neurotransmitters – Overview of Synthesis and Metabolism 1

Definition A chemical released by one neuron that affects another neuron or an effector organ (e.g., muscle, gland, blood vessel)

3 Neurotransmitters Properties of neurotransmitters: 1) synthesized in the presynaptic neuron 2) Localized to vesicles in the presynaptic neuron 3) Released from the presynaptic neuron under physiological conditions 4) Rabidly removed from the synaptic cleft by uptake or degradation 5) Presence of receptor on the post-synaptic neuron. 6) Binding to the receptor elicits a biological response R.E.B, 4MedStudents.com, 2003

4

5 Neurotransmitters found in the nervous system EXCITATORY Acetylcholine Aspartate Dopamine Histamine Norepinephrine Epinephrine Glutamate Serotonin INHIBITORY GABA Glycine

6 Acetylcholine synthesis: In the cholinergic neurons acetylcholine is synthesized from choline. This reaction is activated by cholineacetyltransferase As soon as acetylcholine is synthesized, it is stored within synaptic vesicles.

7 Release of acetylcholine from presynaptic neurons: 1)When the nerve impulse (Action potential) moves down the presynaptic axon to the terminal bulb the change in the membrane action potential causes the opening of voltage gated calcium channels open allowing Ca 2+ ions to pass from the synaptic cleft into the axon bulb. 2) Within the bulb the increase in Ca 2+ concentration causes the synaptic vesicles that contain acetylcholine to fuse with the axonal membrane and open spilling their contents into the synaptic cleft.

8 Binding of acetylcholine to the postsynaptic receptors: The postsynaptic membrane of the receptor dendrite has specific cholinergic receptors toward which the neurotransmitter diffuses. Binding of acetylcholine trigger the opening of ion channels in the postsynaptic membrane initiating action potential that can pass in the next axon. Acetylcholine receptors: Acetylcholine receptors are ion channels receptors made of many subunits arranged in the form [(α2)(β)(γ)(δ)].  When Acetylcholine is not bounded to the receptors, the bulky hydrophobic leu side close the central channels preventing the diffusion of any ions.  Binding of two acetylcholine molecules to the receptors will rotate the subunits in which the smaller polar residues will line the ion channel causing the influx of Na+ into the cell and efflux of K+ resulting in a depolarization of the postsynaptic neuron and the initiation of new action potential.

9 Removal of Acetylcholine from the synaptic cleft: In order to ready the synapse for another impulses: 1) The neurotransmitters, which are released from the synaptic vesicles, are hydrolyzed by enzyme present in the synaptic cleft “Acetylcholinestrase” giving choline, which poorly binds to acetylcholine receptors. Acetylcholine + H2O Choline + H+ acetate 2) The empty synaptic vesicles, which are returned to the axonal terminal bulb by endocytosis, must be filled with acetylecholine.Acetylcholinestrase

10 Structure of AchE Acetylcholinesterase (AchE) is an enzyme, which hydrolyses the neurotransmitter acetylcholine. The active site of AChE is made up of two subsites, both of which are critical to the breakdown of ACh. The anionic site serves to bind a molecule of ACh to the enzyme. Once the ACh is bound, the hydrolytic reaction occurs at a second region of the active site called the esteratic subsite. Here, the ester bond of ACh is broken, releasing acetate and choline. Choline is then immediately taken up again by the high affinity choline uptake system on the presynaptic membrane.

11 Catecholamine Synthesis (Dopamine, Norepinephrine and Epinephrine). 1) First Step: Hydroxylation: In this step: the reaction involves the conversion of tyrosine, oxygen and tetrahydrobiopterin to dopa & dihydrobiopterin. This reaction is catalyzed by the enzyme tyrosine hydroxylase. It is irreversible reaction. 2) Second step: Decarboxylation: In this step: the dopa decaboxylase will catalyze the decaoxylation of dopa to produce dopamine. The deficiency of this enzyme can cause Parkinson’s disease. It is irreversible reaction. The cofactor in this reaction is the PLP (pyridoxal phosphate). In the nerve cells that secrete dopamine as neurotransmitter the pathway ends at this step.

12 3) Third step: Hydroxylation: This reaction is catalyzed by the enzyme dopamine β- hydroxylase. The reactants include dopamine, O 2 and ascorbate (vitamin C). The products are norepinephrine, water and dehydroascorbate. It is an irreversible reaction). The end product in noradrenergic cells is norepinephrine and the pathway ends her. 4) Forth step: Methylation: This reaction is catalyzed by phenylethanolamine N- methyltransferase. Norepinephrine and S-adenosylmethionin (ado-Met) form epinephrine and S-adenosyl homocysteine (ado- Hcy). Catecholamine Synthesis (Dopamine, Norepinephrine and Epinephrine).

13

14 Serotonin synthesis: Serotonin is synthesized from the amino acid Tryptophan. The synthesis of serotonin involve two reactions: 1) 1) Hydroxylation: Tryptophan 5- Hydroxytryptophan The enzyme catalyzes this reaction is Tryptophan Hydroxylase. The Co- factor is Tetrahydrobiopterin, which converted in this reaction to Dihydrobiopterin. 2) 2) Decarboxylation: 5- hydroxytryptophan Serotonin The enzyme is hydroxytryptophan decarboxylase. Serotonin is synthesized in CNS, & Chromaffin cells.

15

16 Break down of serotonin: Serotonin is degraded in two recations 1) Oxidation: 5-hydroxytryptoamine + O2 + H2O 5- Hydroxyinodole-3-acetaldehyde 2) Dehydrogenation 5- Hydroxyinodole-3-acetaldehyde 5-hydroxindole-3-acetate (Anion of 5-hydroxyindoleacetic acid) Monoamine oxidase Aldehyde dehydrogenase

17 Other Neurotransmitters: Neurotransmitter Derived fromEnzyme HistamineHistidineHistidine decarboxylase GABA (γ-Amino butyrate) GlutamateGlutamate decarboxylase Nitric OxideArginineNitric Oxide Synthase

Neurotransmitter Molecule Derived From Site of Synthesis AcetylcholineCholineCNS, parasympathetic nerves Serotonin Serotonin 5-Hydroxytryptamine (5-HT) TryptophanCNS, chromaffin cells of the gut, enteric cells GABAGlutamateCNS HistamineHistidinehypothalamus Epinephrine synthesis pathway Tyrosineadrenal medulla, some CNS cells Norpinephrine synthesis pathway TyrosineCNS, sympathetic nerves Dopamine synthesis pathway TyrosineCNS Nitric oxide, NOArginineCNS, gastrointestinal tract Summary:

The Four Criteria that a Chemical Must Meet to be Called a Neurotransmitter 1. It must be synthesized in the presynaptic neuron - Proof = Biochemical evidence (the biosynthetic enzymes are present in the presynaptic neuron)

The Four Criteria that a Chemical Must Meet to be Called a Neurotransmitter (cont’d) 2. It must be present in the presynaptic neuron and released in adequate quantities to serve as the transmitter at the synapse Proof = Anatomical – the “labeled” transmitter must be present in the presynaptic terminals (e.g., “feed” cell labeled precursors)

The Four Criteria that a Chemical Must Meet to be Called a Neurotransmitter (cont’d) 2. It must be present in the presynaptic neuron and released in adequate quantities to serve as the transmitter at the synapse Proof = Physiological – Collect substance as it is released from the presynaptic neuron and demonstrate that this amount can cause the appropriate postsynaptic response. (NOTE: This has only been done for Ach at the NMJ!)

The Four Criteria that a Chemical Must Meet to be Called a Neurotransmitter (cont’d) 3. When applied exogenously or when its agonists (produce same effect as neurotransmitter) or antagonists (oppose effects of neurotransmitter) are applied to the postsynaptic cell, appropriate responses by the postsynaptic cell must be elicited Proof = Pharmacological - opens appropriate channel, activates appropriate 2 nd messenger system, etc.

The Four Criteria that a Chemical Must Meet to be Called a Neurotransmitter (cont’d) 4. A specific mechanism exists to terminate the presynaptic signal Proof = demonstrate one of the following mechanisms 1. enzymatic breakdown – e.g., Ach: Acetylcholine esterase is synthesized and released by the muscle cell into the basal lamina of the neuromuscular junction. The enzyme cleaves Ach into choline and acetate to terminate the signal. The motor neuron has transporters for choline, used in resynthesis of Ach.

The Four Criteria that a Chemical Must Meet to be Called a Neurotransmitter (cont’d) 4. A specific mechanism exists to terminate the presynaptic signal Proof = demonstrate one of the following mechanisms 2. “presynaptic reuptake” – e.g., Amino acid and biogenic amines use transporters in the presynaptic neuron or surrounding astrocytes to remove signal from synapse.

The Four Criteria that a Chemical Must Meet to be Called a Neurotransmitter (cont’d) 4. A specific mechanism exists to terminate the presynaptic signal Proof = demonstrate one of the following mechanisms 3. diffusion – no longer clear how significant this mechanism is – remember the “tripartite synapse”

The Four Criteria that a Chemical Must Meet to be Called a Neurotransmitter (cont’d) Note how important demonstration that 1) the suspected transmitter is present in/released by the presynaptic neuron in adequate quantities to produce an appropriate postsynaptic response, and, 2) that pharmacological manipulations produce predicted responses are to documenting a chemical’s role as a neurotransmitter. Particularly since very common substances such as glutamate and glycine are purported neurotransmitters

Classes of Neurotransmitters A. Small Molecule Transmitters (“classical transmitters”) – synthesized in cytoplasm from compounds of intermediary metabolism Acetylcholine Biogenic Amines (dopamine, norepinephrine, epinephrine, serotonin, histamine) Amino Acids (gamma-amino butyric acid = GABA, glycine, glutamate) Nitric oxide – small diatomic gas that is also a free radical

Classes of Neurotransmitters B. Neuroactive Peptides – synthesized on rough ER and processed for secretion by the Golgi apparatus 1. Hypothalamic-releasing Hormones 2. Neurohypophyseal Hormones (vasopressin = ADH and oxytocin) 3. Adenohypophyseal Hormones (e.g., Growth hormone, endorphins, melanocyte-stimulating hormone) 4. Gastrointestinal Peptides – (e.g., Gastrin, substance P, insulin, glucagon, vasoactive intestinal peptide) 5. Others – (angiotensin, bradykinin, sleep peptides) HOWEVER, IT IS DEBATEABLE WHETHER THESE ARE NEUROTRANSMITTERS OR A CLASS BY THEMSELVES: HORMONES

HOWEVER, IT IS DEBATEABLE WHETHER THESE ARE NEUROTRANSMITTERS OR A CLASS BY THEMSELVES: HORMONES, because… these do not fit the first 2 criteria of that of neurotransmitter. their actions are far-reaching, rather than just synaptic.

Original Dale’s Law (1950’s) “A mature neuron makes use of the same transmitter substance at all of its synapses” Discovery of peptide transmitters Modified Dale’s Law = “A mature neuron makes use of the same combination of chemical transmitters at all of its synapses.”

Modified Dale’s Law – when co- secretion occurs, it usually involves a small molecule transmitter and one or more peptides Small Molecule Transmitter AcH Norepinephrine Dopamine Epinephrine Serotonin Peptide VIP Somatostatin + enkephalin + neurotensin Cholecystokinin + enkephalin Enkephalin Substance P + TRH