Transporters catecholamine storage

Philippu A. Matthaei H. Transport and storage of catecholamines in vesicles. In Trendelenburg U. Weiner N. eds. Catecholamines I. New York Springer. 1988 1-42. 

The pathway for synthesis of the catecholamines dopamine, noradrenaline and adrenaline, illustrated in Fig. 8.5, was first proposed by Hermann Blaschko in 1939 but was not confirmed until 30 years later. The amino acid /-tyrosine is the primary substrate for this pathway and its hydroxylation, by tyrosine hydroxylase (TH), to /-dihydroxyphenylalanine (/-DOPA) is followed by decarboxylation to form dopamine. These two steps take place in the cytoplasm of catecholaminereleasing neurons. Dopamine is then transported into the storage vesicles where the vesicle-bound enzyme, dopamine-p-hydroxylase (DpH), converts it to noradrenaline (see also Fig. 8.4). It is possible that /-phenylalanine can act as an alternative substrate for the pathway, being converted first to m-tyrosine and then to /-DOPA. TH can bring about both these reactions but the extent to which this happens in vivo is uncertain. In all catecholamine-releasing neurons, transmitter synthesis in the terminals greatly exceeds that in the cell bodies or axons and so it can be inferred... 

Ordinarily, low concentrations of catecholamines are free in the cytosol, where they may be metabolized by enzymes including monoamine oxidase (MAO). Thus, conversion of tyrosine to l-DOPA and l-DOPA to dopamine occurs in the cytosol dopamine then is taken up into the storage vesicles. In norepinephrine-containing neurons, the final P-hydroxylation occurs within the vesicles. In the adrenal gland, norepinephrine is N-methylated by PNMT in the cytoplasm. Epinephrine is then transported back into chromaffin granules for storage. 

Despite the documented efficacy and safety of the psychostimulants, their mechanism of action is not fully understood. Stimulants affect central nervous system (CNS) dopamine (DA) and norepinephrine (NE) pathways crucial in frontal lobe function. The stimulants act by causing release of catecholamines from the DA axons and blocking their reuptake. Methylphenidate releases catecholamines from long-term stores, so its effects can be blocked by pretreatment with reserpine. Amphetamines, on the other hand, release catecholamines from recently formed storage granules near the surface of the presynaptic neuron, so their action is not blocked by reserpine. In addition, the stimulants bind to the DA transporter in striatum (see Figures 2.6 and 2.7) and block the reuptake of both DA and NE. This action reduces the rate that catecholamines are removed from the synapse back into the axon and leads... 

The effect of released norepinephrine wanes quickly, because -90% is transported back into the axoplasm by a specific transport mechanism (norepinephrine transporter, NAT) and then into storage vesicles by the vesicular transporter (neuronal reuptake). The NAT can be inhibited by tricyclic antidepressants and cocaine. Moreover, norepinephrine is taken up by transporters into the effector cells (extraneuronal monoamine transporter, EMT). Part of the norepinephrine undergoing reuptake is enzymatically inactivated to normetanephrine via catecholamine O-methyltransferase (COMT, present in the cytoplasm of postjunctional cells) and to dihydroxymandelic acid via monoamine oxidase (MAO, present in mitochondria of nerve cells and postjunctional cells). 

Storage of catecholamines and serotonm in vesicular granules is facilitated by two vesicular monoamine transporters, both of which are expressed in endocrine cells and one only in neurons. Both transporters have a wide specificity for different monoamine substrates. 

In addition to terminating the actions of released monoamines, the plasma membrane monoamine transporters present at neuronal locations function in sequence with vesicular monoamme transporters to recycle catecholamines for rerelease (Figure 29-3). Thus most of the norepinephrine released and recaptured by sympathetic nerves is sequestered back into storage vesicles, thereby substantially reducing the requirements for synthesis of new transmitter. 

Until recently it was believed that intraneuronal proteins known as serotonin binding proteins (SBP) were involved in the storage, protection, and/or transport of 5-HT (Tamir et al. 1976 Gershon and Tamir 1984) and catecholamines (Jimenez Del Rio et al. 1992,... 

Reserpine lowers BP by depleting norepinephrine from sympathetic nerve endings and blocking transport of norepinephrine into its storage granules. Norepinephrine release into the synapse following nerve stimulation is reduced and results in rednced sympathetic tone, peripheral vascular resistance, and BP. Reserpine also depletes catecholamines from the brain and the myocardinm, which may lead to sedation, depression, and decreased cardiac ontpnt. 

Guanethidine (initial dose 10 mg daily) is indicated in the management of moderate to severe hypertension. It is transported to presynaptic terminals by the catecholamine uptake mechanism, and then slowly displaces norepinephrine from its storage sites to be metabolized presynaptically. 

In addition to synthesis of new transmitter, NE stores are also replenished by transport ofNE previously released to the extracellular fluid by the combined actions of a NE transporter (NET, or uptake 1) that terminates the synaptic actions of released NE and returns NE to the neuronal cytosol, and VMAT-2, the vesicular monoamine transporter, that refills the storage vesicles from the cytosolic pool ofNE ("see below). In the removal ofNE from the synaptic cleft, uptake by the NET is more important than extraneuronal uptake (ENT, uptake 2). The sympathetic nerves as a whole remove -87% of released NE via NET compared with 5% by extraneuronal ENT and 8% via diffusion to the circulation. By contrast, clearance of circulating catecholamines is primarily by nonneuronal mechanisms, with liver and kidney accounting for >60% of the clearance. Because VMAT-2 has a much higher affinity for NE than does the metabolic enzyme, monoamine oxidase, over 70% of recaptured NE is sequestered into storage vesicles. 

Fig. 48.5. Transport of catecholamines into storage vesicles. This is a secondary active transport based on the generation of a proton...

Catecholamine vesicle pump Storage vesicle transporter that pumps amine from cytoplasm into vesicle... 

The PC 12 cell line has also been used to probe the mechanisms of action of various neurologically active drugs. For instance, Sulzer et al. have investigated the effects of amphetamine on exocytosis from PC 12 cells [52], Two distinct mechanisms of action have been proposed (1) exchange diffusion [53] and (2) vesicle depletion [54,55], The exchange-diffusion model involves the attachment of extracellular amphetamine to the dopamine transporter, and the subsequent transport of amphetamine into the cell while dopamine is simultaneously transported out of the cell. This model appears to predominate at low concentrations of applied amphetamine [52]. The second mechanism of action, apparently operational at high doses, involves the depletion of catecholamine from intracellular storage vesicles followed by reverse transport out of the cell [52]. The ampero-metric detection of individual exocytosis events at PC 12 cells provides a unique method to probe this second mechanism. 

Present evidence indicates that the materials of which storage vesicles are composed are synthesized in the nerve cell-body. There is a rapid transport of vesicle material (chromogianins, dopamine-/ -hydroxylase and catecholamines) down the adrenergic nerve axon, at a rate of 5-10 mm/hr. Thus, if a ligature is tied around an adrenergic nerve a rapid accumulation of these materials occurs on the proximal side of the constriction. This transport continues for some time even if tbe nerve is isolated om the cell-body. 

It is probable that some form of mediated transport occurs whenever catecholamines, which are polar compounds, pass through a biological membrane. Thoe are three known processes of this type, which will be described. They are (1) the transport of NA and other amines from the extracellular fluid amoss the neuronal membrane of adrenergic nerves (2) transport of catecholamines from the cytoplasm into storage vesicles (3) transport of catecholamines from the extracellular fluid into the cytoplasm of non-neuronal cells. 

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