Splenic Nerve Research Articles

Axoplasmic transport of muscarinic receptors.

The reality of axoplasmic transport is widely accepted; various neutrotransmitters, enzymes, labelled proteins and peptides are known to move rapidly along the axons of different nerve fibres. In the terminals of sympathetic nerves, noradrenaline release is controlled by various regulatory mechanisms which imply the occurrence of presynaptic receptors. In this regard, there is considerable indirect physiological evidence for the existence of muscarinic cholinergic receptors in the sympathetic nerve endings; the stimulation by acetylcholine of such presynaptic receptors elicits an inhibitory effect on noradrenaline release. We not provide direct biochemical evidence for the occurrence in dog splenic nerve of muscarinic receptors which seem to move along the axon as suggested by their rapid accumulation on either side of a ligature.

Dopamine β‐hydroxylase distribution in density gradients: Physiological and artefactual implications

Knowledge of the vesicular origin of circulating dopamine beta-hydroxylase (DbetaH) is indispensable for any attempts to explain the parallelism or lack of it between circulating enzyme and catecholamines as they may relate to physiological stress, forms of hypertension, neurological disorders, and the response to pharmacological agents. The present study represents an effort to evaluate and to place in proper perspective data based on the DbetaH activity found in the region of the light vesicle peak of noradrenaline (NA), which is used as a quantitative measure of a population of small terminal vesicles. Distributions of vesicles and subvesicular components are compared with DbetaH and NA in sucrose-D2O density gradients used to prepare relatively pure fractions of large dense cored vesicles (LDV) from bovine splenic nerve. Although NA in sedimentable particles of the light vesicle peak is likely to be a valid measure of a small vesicle population, the following is demonstrated: (1) A substantial fraction (25%-37%) of the total sedimentable DbetaH activity can be proven to distribute in the region of the light vesicle peak from a tissue with an insignificant small vesicle population. Based on studies of vesicles from sequential nerve segments, this enzyme activity probably corresponds to a population of "immature" LDV which are undergoing axoplasmic transport and have not synthesized their full complement of transmitter. (2) Physical lysis which depletes the matrix of LDV causes redistribution of DbetaH activity from the heavy vesicle peak into the region of the light vesicle peak. Analogously, DbetaH associated with exocytosed LDV and retrograde transport particles is also likely to contaminate the region of the light vesicle peak. (3) Based on available data, it can be calculated that each small dense cored vesicle could contain only 0.1-0.5 molecules of DbetaH and that a contamination of only 0.016% LDV can account for all of the DbetaH reported to occur in the light vesicle peak of normal rat vas deferens preparations.

Catecholamine‐rich cells and varicosities in bovine splenic nerve, vesicle contents and evidence for exocytosis

The bovine splenic nerve trunk contains mast cells, ganglion cells, small intensely fluorescent (SIF) cells, and varicosities which exhibit a brilliant fluorescence characteristic for noradrenaline (NA) and dopamine (DA) after formaldehyde exposure. All these catecholamine-rich structures could contribute particles to isolated nerve vesicle fractions. Mast cells are recognized ultrastructurally by their large (300-800 nm) dense granules. SIF cells may be represented by cells and processes containing dense cored vesicles (120-140 nm) which are larger than the typical vesicles in axons and terminals. Terminal-like areas with typical large dense cored vesicles (LDV, 75 nm) and small dense cored vesicles (SDV, 45-55 nm) probably correspond to the fluorescent varicosities. The LDV constitute about 40% of all vesicles in terminal-like areas and terminals. Their staining properties indicate the presence of protein, phospholipids, and ATP. Tyramine depletes NA without loss of matrix density. The LDV can fuse with the terminal membrane, and released material outside omega profiles is interpreted to depict exocytosis. Large and small vesicles are easily distinguished from the very large mast cell granules and the moderately dense Schwann cell vesicles. Neither appear to contaminate the LDV fractions but the latter may contain a small population of SIF cell vesicles. Golgi vesicles from the Schwann cells mainly occur in the lighter zones of the gradient.

Effect of splenic nerve stimulation on the contents of noradrenaline, ATP and sulphomucopolysaccharides in noradrenergic vesicle fractions from the cat spleen

In order to assess the effect of splenic nerve stimulation on the contents of noradrenaline, ATP and 35S-labelled compounds in noradrenergic vesicles from the spleen, experiments were carried out on the cat spleen in situ, its frontal portion being used as control. Particulate fractions containing noradrenergic vesicles were isolated on Ficoll gradient centrifugation of homogenates from both portions of the spleen. The cats received phenoxybenzamine (10 mg/kg) and imipramine (5 mg/kg) to inhibit the contractive response of the spleen to nerve stimulation and the reuptake of noradrenaline (NA) into nerve terminals. Administration of the drugs did not significantly affect the levels of NA, ATP and 35S (presumably in sulphomucopolysaccarides, SMPSs). In response to prolonged stimulation of the splenic nerve a significant loss of NA from noradrenergic vesicles occurred, whereas levels of ATP and 35S-labelled compounds remained unchanged. The results indicate that a discharge of NA in response to nerve stimulation is not accompanied by simultaneous discharge of ATP and SMPSs.

Effects of adenosine on neurally mediated norepinephrine release from the cat spleen

The effects of adenosine on the release of 3H-norephrine ( 3H-NE) from the isolated, perfused cat spleen consequent to nerve stimulation were evaluated. Electrical stimulation of the splenic nerve (5 Hz/100 impulses total) caused a release of 3H-NE and a rise in perfusion pressure. Adenosine added to the perfusion fluid (final concentrations 1 × 10 −6, 1 × 10 −5, and 1 × 10 −4 M) significantly reduced the pressure response elicited by nerve stimulation. In addition, adenosine (1 × 10 −4 M) slightly increased the release of total 3H consequent to nerve stimulation. Theophylline (1 × 10 −4 M) produced both a slight increase in total 3H and a diminished pressure response. The effects of adenosine were effectively antagonized by this concentration of theophylline. Neither substance had any effect on the spontaneous release of total 3H. Adenosine (1 × 10 −4 M) also antagonized the pressure response elicited by perfusion of NE.

Choline acetyltransferase in the equine spleen

1. 1. Choline acetyltransferase (ChAc) has been investigated in the spleens of the horse ( Equus cahallus) and donkey ( Equus asinus) and found to be present in amounts ranging from 0.34–6.86 μmol of ACh synthesized g −1 hr −1. Spleens of other ungulates investigated contained much less than this. 2. 2. ACh content of the spleens studied corresponded with these findings. 3. 3. ChAc was not detected in either the splenic nerve nor in subcellular fractions which could be identified as parts of nerve endings. 4. 4. When splenic sections were stained for acetylcholinesterase very few nerve fibres could be found. On the assumption that cholinergic nerve fibres should be so stained there appear to be far too few such fibres to account for the ChAc found. 5. 5. No evidence was found for a powerful inhibitor to ACh synthesis in horse spleen as previously reported. 6. 6. The results are discussed in relation to possible roles for ACh in the post-ganglionic sympathetic release of noradrenaline.

The formation of actin-containing gels by extracts of bovine splenic nerve

Extracts prepared from bovine splenic nerve at low ionic strength were concentrated by vacuum dialysis. When incubated with magnesium, adenosine triphosphate and the calcium chelating agent ethyleneglycol-bis(β-amino-ethyl)N,N'-tetra-acetic acid at 37°C for 1 h, the concentrated extracts formed strong, invertible gels. Guanosine triphosphate but not adenyl methylene diphosphate could substitute for adenosine triphosphate. Calcium ions and KCl above 0.1 M inhibited gelation. Gelation did not occur at 0°C. Gelation was prevented by cytochalasin and phalloidin but not by colchicine, podophyllotoxin or vinblastine.Sodium dodecyl sulphate electrophoresis of proteins sedimented from the invertible gels showed polypeptides which co-migrated with actin, α and β tubulin, bovine plasma albumin and myosin. Amongst the other polypeptides there was one with a molecular weight of about 215,000 and a doublet corresponding in mobility with the microtubule accessory proteins of high molecular weight.It is concluded that extracts of splenic nerve are able to form gels stabilized by fibres of actin which might be crosslinked by the polypeptides of molecular weight 215,000. The origin of actin found in extracts of splenic nerve is discussed.

The differential effect of cooling on the responses of splenic capsular and vascular smooth muscle to nerve stimulation and noradrenaline.

The responses of the capsular and vascular smooth muscle of the dog's spleen to splenic nerve stimulation and to infused noradrenaline have been studied in the isolated, blood-perfused preparation at 37 degrees C, at 27 degrees C and again after rewarming to 37 degrees C. It was found that cooling per se had no effect on perfusion pressure but reduced splenic arterial blood flow, and caused no appreciable alteration in spleen volume. The increase in splenic flow resistance in response to nerve stimulation and noradrenaline was significantly greater at 27 degrees C than at 37 degrees C, but the concomitant reduction in spleen volume was significantly reduced. The enhanced effect of splenic nerve stimulation and noradrenaline on splenic flow resistance is discussed in terms of the relative contributions of an increased smooth muscle response and increased blood viscosity. The different effects of cooling on the responses of splenic vascular and capsular smooth muscle to nerve stimulation and noradrenaline are discussed in the context of the effect of cooling on other vascular and non-vascular smooth muscle.

Evidence that most of the dopamine β-hydroxylase is not membrane bound in purified large dense cored noradrenergic vesicles

The compartmentalization of dopamine β-hydroxylase was studied in a purified fraction of large, dense-cored noradrenergic vesicles from bovine splenic nerve. A correlative biochemical and morphological approach was used to evaluate physical methods designed to produce more efficient vesicle lysis than has been accomplished previously with freeze-thawing, hypo-osmotic shocks or incubation at 37°C, all of which are relatively ineffective. Of the seven purely physical techniques tested, only those which subjected the vesicles to high pressures followed by rapid decompression produced effective lysis, unmasking of latent enzyme activity, and release of enzyme into the soluble phase. Based on results with the French Press, the best estimate for the percentage of dopamine β-hydroxylase which can be released into the soluble phase is 55–70% of the total enzyme activity produced by the addition of 0.05% v/v Triton X-100. This includes two-thirds of the enzyme which was originally in a latent form. The enzyme is considered to occur primarily, if not entirely, in a granular complex in the vesicle matrix. About one-third is thought to be oriented specifically at the inner surface of the vesicle membrane and is active enzymatically. Physical rupture of the vesicles causes redistribution of partially depleted vesicles and free matrix granules containing dopamine β-hydroxylase activity into less dense zones of a sucrose-D 2O density gradient. After French Press treatment, 30% of the enzyme lost from the heavy vesicle peak is found in the region of the light vesicle peak which is thought to be the equilibrium density of the smaller vesicles characteristic of nerve terminals. The data indicate that much more dopamine β-hydroxylase is potentially available for release from the large, dense-cored vesicles upon nerve stimulation than previously has been believed. This will affect interpretations of exocytotic release from the two vesicle populations in noradrenergic varicosities and will have a bearing on the origin of circulating dopamine β-hydroxylase in response to physiological stress and pharmacological intervention, as well as in neurological diseases.

The effects of labetalol (AH 5158) on metabolism of 3H(−)-noradrenaline released from the cat spleen by nerve stimulation

The competitive α and β adrenoceptor antagonist labetalol, in concentrations up to 10 −4 M, produced a dose dependent increase in overflow of 3H and [ 3H]noradrenaline in the isolated blood perfused cat spleen following stimulation of the splenic nerves at a frequency of 10 Hz. Labetalol had no effect on the pattern of overflow of label following stimulation. In experiments in which the metabolism of [ 3H]noradrenaline released on nerve stimulation was examined, labetalol produced a concentration dependent increase in the percentage of [ 3H]noradrenaline and a decrease in the percentage of [ 3H]DOPEG in the venous blood following nerve stimulation. Production of [ 3H]COMT metabolites and [ 3H]DOMA was not affected. It is suggested that in the isolated blood perfused cat spleen labetalol produces the elevation of overflow and effects on noradrenaline metabolism by inhibition of neuronal uptake of noradrenaline. The drug has no detectable effects on the enzymes MAO or COMT or on extraneuronal uptake.

Latency of dopamine β-hydroxylase in purified noradrenergic vesicles

Purified large dense core noradrenergic vesicles isolated from bovine splenic nerve were investigated to determine the percentage of total dopamine β-hydroxylase which existed in a latent or masked form. The effects of the nonionic detergent Triton X-100 in concentrations between 0.0025 and 0.1% were studied. At 0.01%, Triton produced a maximum three-fold increase in accessible dopamine β-hydroxylase activity to reach 0.64μmol octopamine per mg protein per min at 37°C, but released only a third of the enzyme into the soluble phase. About 90% of the noradrenaline, 42% of the phospholipids and 17% of the vesicle protein were released. At 0.05%, Triton produced the same maximum enzyme activity with complete solubilization of the enzyme. All the noradrenaline, 90% of the phospholipids, and 48% of the vesicle protein were released into the soluble phase. Ultrastructural examination of the treated vesicles revealed only partial lysis with 0.01% Triton, while 0.05% produced essentially complete disruption of vesicles into membrane fragments and fine granular matrix contents. Equilibrium centrifugation of the control and treated vesicles on a second sucrose-D 2O density gradient showed that emptied and partially depleted vesicles and released matrix granules redistributed from the heavy vesicle peak to less dense regions of the gradient, including the zone of the light vesicle peak. It is proposed that the latency of dopamine β-hydroxylase activity in this noradrenergic vesicle fraction is best explained by the occurrence of the enzyme as an inaccessible complex in the vesicle matrix which may include phospholipid.

The effects of labetalol (AH 5158) on adrenergic transmission in the cat spleen.

1. The competitive alpha- and beta-adrenoceptor blocking agent labetalol, in concentrations up to 10(-4) M, produced dose-dependent increases in transmitter overflow from the isolated blood perfused spleen of the cat following nerve stimulation at 10 and 30 Hz. 2. At concentrations above 10(-4) M labetol produced a pronounced decrease in transmitter overflow. 3. Labetalol (1.5 X 10(-4) M) increased the recovery of 3H label in the venous blood following the close-arterial infusion of [3H]-(-)-noradrenaline indicating that the drug inhibits uptake of the amine. 4. Both labetalol (3.8 X 10(-5) M) and piperoxan (7.4 X 10(-6) M) produced parallel shifts to the right of the dose-response curves to noradrenaline and oxymetazoline in isolated strips of cat splenic capsule. In this preparation both drugs acted as competitive postsynaptic alpha-adrenoceptor blocking agents. 5. Labetalol (3.3 X 10(-5) M) increased the transmitter overflow following stimulation of the splenic nerves with 200 impulses at 10 Hz. The overflow could be further increased by subsequent addition of piperoxan (7.2 X 10(-6 M). Piperoxan (5.7 X 10(-6) M) alone produced a marked increase in transmitter overflow which could be further increased by subsequent addition of desmethylimipramine (DMI; 3.2 X 10(-5) M). Cocaine (1.5 X 10(-5) M) or DMI (5.4 X 10(-5 M) produced a small increase in transmitter overflow which was not further increased by addition of labetalol (2.8 X 10(-5) M). 6. Labetalol produced a biphasic effect on the responses of the isolated blood perfused spleen of the cat to nerve stimulation. With low doses (up to 10(-4) M) vascular responses were potentiated and with high doses (greater than 10(-4) M) inhibited. The potentiation was related to uptake blockade and the inhibition to decreased transmitter overflow and postsynaptic alpha-adrenoceptor blockade. 7. Labetalol appears to act as a postsynaptic alpha-adrenoceptor antagonist in the isolated blood perfused spleen of the cat with little effect on presynaptic alpha-adrenoceptors. The moderate elevation of transmitter overflow by the drug is related to the inhibitory effect of the drug on neuronal uptake rather than on presynaptic alpha-adrenoceptors.

Biochemical observations on the formation of small noradrenergic vesicles in the splenic nerve of the dog

Splenic nerve terminals contain noradrenergic vesicles which can be classified as small and large on the basis of both morphological and biochemical experiments. The origin of the large vesicles is likely to be the cell body and they are probably supplied to the nerve terminals by axonal transport. The origin of the small noradrenergic vesicles is still obscure so we have tested, using centrifugation techniques, the hypothesis that they are formed from the large noradrenergic vesicles during secretory activity in the nerve endings. Normally the small noradrenergic vesicles in resting splenic nerve terminals contain very little dopamine β-hydroxylase activity, as distinct from the large vesicles. However, after stimulation in vitro of the perfused spleen there is an appreciable amount of the enzyme associated with the small particles. These newly formed small vesicles are able to concentrate exogenous noradrenaline and their formation is dependent upon release of secretory products from the large vesicles as shown by their absence if the splenic nerve of a spleen perfused with a Ca 2+ -free solution is stimulated. A balance sheet has been drawn up which suggests that membrane-bound dopamine β-hydroxylase lost from the large noradrenergic vesicles upon stimulation is almost quantitatively recovered in the small noradrenergic vesicles. It is suggested that at least three types of noradrenaline storage particles exist in splenic nerve terminals (i) large vesicles which contain noradrenaline and dopamine β-hydroxylase and that can give rise to (ii) small vesicles which also contain both constituents and, (iii) vesicles which are virtually devoid of dopamine β-hydroxylase activity but which can store noradrenaline and have an origin quite different from those of the other two types.

Effect of surugatoxin on celiac ganglia in cats (author's transl)

Pharmacological properties of surugatoxin (SGTX), the toxic principle from Japanese ivory mollusc (Babylonia japonica), have been studied. In the present paper, the effect of SGTX on celiac ganglia in cats was investigated by measuring blood pressure from femoral artery and regional blood flow in liver (LBF) and spleen (SBF) of anaesthetized cats. SGTX(50 nmol/kg i.v.) markedly inhibited both the hypertension and changes of LBF and SBF in response to splanchnic (preganglionic) nerve stimulation. On the other hand, it did not inhibit the changes of SBF by splenic(postganglionic) nerve stimulation. The hypertension and changes in LBF and SBF in response to DMPP(94.2 nmol/kg i.v.) were abolished by SGTX(50 nmol/kg i.v.), whereas those to McN-A-343 (94.8 nmol/kg i.v.) were not. SGTX usually enhanced the pressor response to McN-A-343. These results indicate that SGTX has a blocking effect on the nicotinic receptor in celiac ganglia as well as in superior cervical ganglia of cats and rats.

Preliminary estimates of the dopamine beta-hydroxylase content and activity in purified noradrenergic vesicles.

Abstract—The dopamine β‐hydroxylase (DβH) content and activity of large dense‐core noradrenergic vesicles purified from bovine splenic nerve were determined using two assay procedures : enzymic activity expressed in Units per mg protein and homospecific activity based on radioimmunoassay expressed in Units per mg DβH antigen. Approximately two‐thirds of the total enzyme activity is latent in these vesicles, even after various treatments designed to compromise vesicle integrity. DβH can be completely unmasked by brief treatment with 0.01‐0.05% Triton X‐100 and activity increases from 0.20 to 0.64 Units per mg vesicle protein. Calculations based on both assay methods suggested that an average of 7% (range 3‐15%) of the total vesicle protein was DβH and that the average vesicle contained about 4 molecules of enzyme (range 2‐9 molecules). The estimated homospecific activities indicated an average of 25 and 50% (range 18‐72%) of the vesicle enzyme was inactive in the various samples using the two antibodies. The vesicle can synthesize up to 30 molecules of noradrenaline/s per molecule of DβH at near optimal substrate concentration, and 60‐270 molecules of norepinephrine/s per vesicle. The assumptions used in the various calculations were critically analyzed and, based on the methods employed, it is tentatively considered to be unlikely that there could be more than 5‐12 molecules of DβH per vesicle. The possibility that circulating DβH originates primarily, if not exclusively, from the large dense‐core vesicle type is considered and the functional implications of the data support the concept of vesicle reuse during several cycles of exocytosis involving a quantal size equal to a fraction of the vesicle transmitter content.

Release of noradrenaline from the cat spleen by nerve stimulation and potassium.

1. Release of noradrenaline from the perfused cat spleen, or from isolated spleen slices, in response to prolonged nerve stimulation or maintained depolarization by potassium was measured. 2. Prolonged stimulation of the splenic nerves at 2, 10 and 30 Hz for 10 min evoked release, which was maximum during the first 2 min, and then declined during the remaining period of stimulation. When noradrenaline release was induced by high potassium from the perfused spleen or from isolated slices, it followed a similar time course to nerve stimulation. Similar results were obtained from phenoxybenzamine-treated spleens, using both modes of stimulation. 3. Stimulation of the splenic nerves in calcium-free Krebs solution did not release noradrenaline. If calcium was introduced at a later stage during stimulation, the release was markedly diminished. In phenoxybenzamine- or phentolamine-treated spleens, stimulation of the nerves in the presence of calcium evoked a secretory response which was comparable to the one produced by introduction of calcium after a few minutes of nerve stimulation. 4. Simultaneous application of calcium plus high potassium always produced a much greater secretion of noradrenaline than application of calcium after a few minutes of potassium depolarization. Release of noradrenaline by potassium from phenoxybenzamine-treated spleens was also much greater if calcium and potassium were added simultaneously than addition of calcium after a few minutes of potassium depolarization. 5. In the presence of maintained depolarization by potasssium, tyramine was effective in causing release of noradrenaline.

Relation of efferent impulse activity in splenic nerve to reflexly induced reactions of resistance and capacitance vessels to spleen.

Constrictory responses of splenic resistance vessels arising under pressor reflexes were abolished by hexonium (2 mg/kg) as well as the high amplitude (above 15 muV) impulses in sympathetic splenic nerves. Constrictory and dilatory responses of splenic capacitance vessels were preserved after administration of the same dose of hexonium and correlated as to the directivity with the changes of the low amplitude (15 muV and lower) impulsation in the splenic nerve.

Gel electrophoresis of soluble and insoluble proteins of noradrenergic vesicles from ox splenic nerve: A comparison with proteins of adrenal chromaffin granules

Polyacrylamide gel electrophoresis of a soluble extract of purified large dense-cored vesicles (noradrenergic vesicles) from ox splenic nerve revealed 13 proteins, 7 of which had mobilities close to those of seven of the soluble proteins in adrenal chromaffin granules. Two of these proteins had the same mobilities as dopamine β-hydroxylase and chromogranin A, respectively. The relative staining intensities of the latter two proteins were different: in noradrenergic vesicles, there was more dopamine β-hydroxylase than chromogranin A; whereas in chromaffin granules, chromogranin A was the major protein. Gel electrophoresis of the water-insoluble proteins of noradrenergic vesicles, dissolved in sodium dodecylsulphate, revealed 5 proteins, 5 of which had mobilities close to those of 5 proteins present in the membranes of chromaffin granules. Three of these proteins had mobilities similar to those of dopamine β-hydroxylase, chromogranin A and chromomembrin B, respectively. One of the proteins was probably serum albumin, which was also present as a contaminant in the soluble extract of noradrenergic vesicles. These findings are consistent with earlier studies in which dopamine β-hydroxylase, chromogranin A and chromomembrin B have been identified as constituents of noradrenergic vesicles by enzymatic or immunochemical assay methods. They also indicate further qualitative similarities between the noradrenergic vesicle and the chromaffin granule, in that a total of 7 soluble and 5 insoluble proteins might be common to both particles. However, gel electrophoresis also confirms that quantitative differences exist between the relative proportions of the soluble proteins and shows that dopamine β-hydroxylase is the major soluble protein of the noradrenergic vesicles isolated from splenic nerve trunks.

Identification of the mucopolysaccharides in catecholamine-containing subcellular particel fractions from various rat, cat and ox tissues.

Subcellular fractions containing adrenergic vesicles were obtained from bovine adrenal medulla and splenic nerve, rat and cat heart, and rat spleen. The fractions were washed free from lipids, digested with papain and the mucopolysaccharides (MPSs) then precipitated from the supernatant with ethanolic potassium acetate. The mpsswere identified by several different methods--microelectrophoresis on cellulose acetate in various media, cellulose column chromatography using elution solvents of increasing ionic strength, and treatment with specific enzymes followed by electrophoresis or column chromatography. The MPS precipitate from all the sources investigated contained dermatan sulphate or a dermatan sulphate-chondroitin sulphate hybrid. in addition, the precipitate from rat spleen was found to contain chondroitin sulphate. Heparan sulphate was found in the precipitates from rat heart and spleen and hyaluronic acid in that from bovine splenic nerve. The finding of sulphomucopolysaccharides in the adrenergic vesicles, probably in a complex with protein, raises the question of the functional significance of such complexes. They might, by analogy with the ion-exchange function of the heparin-protein complex in mast cell granules, play a role in the storage and release of the amines.

Biochemical correlates for the pharmacological effects of L(+)-Isomers and β-desoxy-sympathomimetic amines

The pharmacological effects of metabolically stable χ-methylated indirectly acting β-hydroxylated l(+)-isomers and the corresponding β-desoxy-derivalives of phenolic and nonphenolic sympathomimctic amines were investigated. On the isolated, superfused rat vas deferens, preloaded with [ 3H]norepinephrine ( 3H-NE). the contraction and the efflux of tritium from the tissue by the drugs were studied. It was observed that a given phenolic amine can block the effects of other phenolic amines. For example. l(+)-cobefrin can block the effect of (+)-desoxycobefrin or tyramine. A nonphenolic amine. however, blocks other nonphenolic amines but not the effects of phenolic amines. For example. l(+)-ψ-ephedrine can block the effect of (+)-desoxyephedrine but not that of tyramine. The efflux of tritium parallels the pharmacologic effect. It is implied that phenolic and nonphenolic amines utilize different transport systems at the adrenergic nerve terminal. Other consistent findings have been that the desoxy-derivative of either phenolic or nonphenolic amines always produces a greater pharmacologic effect than the corresponding l(+)-isomer. Since their ability to block uptake of 3H-NE is the same. as compared to the l(+)-form, the desoxy-derivative must enter the adrenergic neurone faster and/or displace more transmitter to produce its greater pharmacologic effect. Additional studies with isolated bovine splenic nerve granules indicate that the less effective l(+)-isomers do not release NE from storage vesicles, whereas desoxy-analogs release considerable quantities of NE from these sites. The implications are that the desoxy-form may release NE from both cytoplasmic and vesicular stores, while the l(+)-isomer may release NE only from an extravesicular site.