L. Stjärne

Site(s) and ionic basis of α‐autoinhibition and facilitation of [3H]noradrenaline secretion in guinea‐pig vas deferens

1. Mechanisms controlling the secretion of [3H]noradrenaline from the noradrenergic nerves of guinea‐pig isolated vas deferens, prelabelled by incubation with [3H]noradrenaline, were studied using (a) different modes of (extramural or transmural) electrical nerve stimulation (a total of 300 shocks of varying strength, and a duration of 2 msec) at 1‐30 Hz, or (b) depolarizing concentrations of K+ (60‐110 m m).

Localization of different steps in noradrenaline synthesis to different fractions of a bovine splenic nerve homogenate.

Abstract Experiments were carried out to determine to what extent the synthesis of noradrenaline in bovine splenic nerve tissue can proceed in the high-speed sedimentable particulate fraction that contains the specific noradrenaline storage vesicles. Various fractions of a homogenate of this tissue were incubated in a potassium phosphate medium together with radioactively labeled precursors of noradrenaline. The first two steps in the synthesis, the ring hydroxylation of tyrosine and the decarboxylation of dopa, were shown not to require the presence of this particulate fraction. On the other hand, this fraction was an absolute prerequisite for the last step, the β-hydroxylation of dopamine. No significant formation of catechol compounds from tyrosine could be demonstrated on incubation with this particulate fraction after it had been washed and resuspended in potassium phosphate. Some dopa decarboxylase activity remained in the particulate fraction even after washing. The results support the view that in this tissue only one enzyme involved in noradrenaline synthesis, dopamine β-hydroxylase, is truly located in the above-mentioned particles, possibly inside the membrane of the noradrenaline-storing vesicles. Tyrosine hydroxylase and dopa decarboxylase appear to be located outside these particles. Dopa decarboxylase shows some association with the particulate fraction studied, and might be, to some extent, bound to the outer aspect of the same particles which contain the β-hydroxylase, or could just be accumulated in the perivesicular parts of the probably highly organized axoplasm.

Acceleration of noradrenaline biosynthesis by nerve stimulation

Abstract Prolonged preganglionic stimulation of the isolated hypogastric nerveas deferens preparation resulted in a 3-fold increase in the amount of NA synthesized from exogenous tyrosine. This increase was not the result of an increased effector organ activity since the increase was still apparent even after blockade of the effector organ response with Hydergine®. These results indicate that the isolated vas deferens preparation is capable of synthesizing NA from exogenous tyrosine and that local NA synthesis is in some way regulated by nervous activity and is not a steady state phenomenon independent of impulse traffic.

Acceleration of noradrenaline turnover in the mouse heart by cold exposure

Abstract The rate of disappearance of tritiated noradrenaline (NA) from the heart has been used to estimate the turnover of NA in this tissue (1). Although these two parameters were not regarded as identical, it was claimed that any change in the rate of NA turnover would be reflected in the slope of the tritiated NA disappearance curve. The conclusion reached by these authors was that the NA turnover rate in the mouse heart was constant, independent of nerve activity and corresponded to a NA synthesis rate of about 0.12 μg/g/hour. In the present experiments the question whether the NA synthesis rate in the mouse heart can be accelerated by increased nerve activity was studied by comparing the slopes of the tritiated NA disappearance curve in hearts of mice kept in the cold with that in hearts from control mice kept at room temperature.

Basic Mechanisms and Local Feedback Control of Secretion of Adrenergic and Cholinergic Neurotransmitters

The main aim in this chapter is to review recent evidence indicating that adrenergic and cholinergic neurotransmitter secretion may be subject to local feedback control. To understand the nature of the control mechanism, it is obviously essential to have a realistic picture of the controlled function. Thus before going into the evidence concerning physiological regulation of neurotransmitter secretion, certain crucial aspects of the available information concerning the basic processes involved in adrenergic and cholinergic transmitter secretion will be discussed. The intention is not to extensively review the literature in this vast field, but to point out what seem to be inconsistencies and discrepancies in current concepts concerning the basic processes involved in neurotransmitter secretion.

The influence of 8-Br 3′,5′-cyclic nucleotide analogs and of inhibitors of 3′,5′-cyclic nucleotide phosphodiesterase, on noradrenaline secretion and neuromuscular transmission in guinea-pig vas deferens

SummaryThe effects of 3′,5′-cyclic nucleotide phosphodiesterase (PDE) inhibitors and of 8-Br 3′,5′-cyclic nucleotide analogs on nerve-muscle transmission were studied in the guinea-pig vas deferens preincubated with 3H-noradrenaline.8-Br cyclic AMP and the PDE inhibitors 3-isobutyl-1-methylxanthine (IBMX) and 3-propionyl-4-hydrazinopyrazolopyridine (SQ 20006) enhanced the secretion of 3H-NA evoked by transmural nerve stimulation. 8-Br cyclic GMP was without effect in this respect.The muscle contraction evoked by transmural nerve stimulation, high potassium or by application of exogenous noradrenaline was depressed by IBMX and SQ 20006. The contraction evoked by transmural nerve stimulation was enhanced by 8-Br cyclic AMP and depressed by 8-Br cyclic GMP.These findings suggest differential involvement of 3′,5′-adenosine- and guanosine-cyclic nucleotides in excitation-secretion-coupling in the noradrenergic sympathetic nerves, and in excitation-contraction-coupling in the smooth muscle, of guinea-pig vas deferens.

Michaelis-Menten kinetics of secretion of sympathetic neurotransmitter as a function of external calcium: Effect of graded alpha-adrenoceptor blockade

SummaryThe curve describing the calcium dependence of nerve stimulation induced secretion of tritium labelled noradrenaline (3H-NA) from isolated guinea-pig vas deferens was sigmoid in shape. Graded alpha-adrenoceptor blockade dose dependently moved the curve to the left and shifted it into a rectangular hyperbola. A double reciprocal plot of the fractional secretion of 3H-NA against the calcium concentration in the medium, during disinhibition of the secretory mechanism by alpha-adrenoceptor blockade, yielded a straight line. This indicates that the disinhibited secretory mechanism is basically a simple function of the calcium concentration in the medium, obeying classical Michaelis-Menten kinetics, and normally proceeding at a calcium concentration close to that which gives half maximal secretory rate.

Electrophysiological and electrochemical analysis of the secretion of ATP and noradrenaline from the sympathetic nerves in rat tail artery: effects of α2-adrenoceptor agonists and antagonists and noradrenaline reuptake blockers

SummaryThe aim of this study was to investigate whether or not nerve impulses release ATP and noradrenaline in parallel from the sympathetic nerve terminals of the rat tail artery. The extracellularly recorded excitatory junction current (EJC) was used to study, pulse by pulse, the release of ATP. An electrochemical method was used to study online the nerve stimulation-induced rise in the extracellular concentration of endogenous noradrenaline at the probe, a carbon fibre electrode (CF). This parameter, which does not directly represent noradrenaline release, but reflects release minus clearance, has been termed Δ[NA]CF. The effects of a number of pharmacological agents on the EJCs were examined both at 0.1 and 2 Hz, and the effects on the EJC response to 100 pulses at 2 Hz compared with that on the Δ[NA]CF response. Clonidine and xylazine were used as α2-agonists, yohimbine and idazoxan as α2-antagonists and desipramine and cocaine as blockers of noradrenaline reuptake. Most of these agents had unwanted side effects, especially at higher concentrations. However, clonidine and xylazine depressed at lower concentrations the EJC and Δ[NA]CF responses to about the same extent; these effects were partially or completely reversed by yohimbine. Yohimbine or idazoxan did not affect the EJCs at 0.1 Hz but enhanced the EJC and Δ[NA]CF responses to 100 pulses at 2 Hz to the same extent. All effects of desipramine (1 μM) seemed explainable as a result of block of noradrenaline reuptake, while cocaine (10 μM) in addition exerted an ‘unspecific’ depressant (probably local anesthetic) effect. Under control conditions, both agents depressed the EJC but dramatically enhanced the Δ[NA]CF response to 100 pulses at 2 Hz. Addition of yohimbine prevented the depressant effect of desipramine on the EJCs completely and reduced that of cocaine, but increased their effects on the Δ[NA]CF response. These results are compatible with the view that ATP and noradrenaline are released in parallel from the sympathetic nerve terminals of this tissue. The different, and under some conditions even opposite, effects of desipramine or cocaine on the EJC and Δ[NA]CF responses are explainable in terms of the known post-secretory effects of these agents.

On the secretory activity of single varicosities in the sympathetic nerves innervating the rat tail artery.

1. Nerve terminal impulses (NTIs) and spontaneous or stimulus‐evoked excitatory junction currents (SEJCs or EJCs), reflecting secretion of transmitter quanta from release sites in the sympathetic nerves of rat tail artery, were recorded by extracellular electrodes. 2. The release of transmitter quanta from single varicosities was analysed on a pulse‐by‐pulse basis. 3. Since the SEJCs were tetrodotoxin‐resistant, and hence probably caused by single quanta, they were employed to analyse the quantal content of EJCs. 4. In the majority of recordings, EJCs were large compared to SEJCs from the same attachment, and preceded by prominent NTIs. This type of activity appeared to reflect simultaneous activation of several nerve fibres and numerous varicosities. 5. By focal stimulation, it was usually possible to improve the resolution by examining spots in which a large proportion of the suprathreshold stimuli failed to cause EJCs. Here, averaged NTIs preceding large EJCs were indistinguishable from averaged NTIs not followed by EJCs. Thus, failure of invasion by the nerve impulse was not a cause of the frequent secretory failure. 6. In these attachments the amplitude distribution of nerve stimulus‐evoked EJCs was similar to that of the SEJCs and many individual EJCs could be matched in amplitude and time course by SEJCs. Thus, transmitter secretion from these sympathetic nerve varicosities seems to be basically monoquantal. 7. Under conditions when all EJCs were smaller than or equal to the largest SEJCs some characteristic EJC profiles appeared only a few times in response to several hundred suprathreshold stimuli at low frequency (0.5‐1 Hz). Using tentatively these EJCs as ‘fingerprints’ of single quanta from particular release sites, the probability for activation of individual release sites ranges from 0.002 to 0.02.

Selectivity for catecholamines of presynaptic alpha-receptors involved in feedback control of sympathetic neurotransmitter secretion in guinea-pig vas deferens

SummaryThe aim of the study was to quantitatively compare the relative affinities of noradrenaline, adrenaline, dopamine and isoprenaline for the, probably neural, receptors mediating feedback control of sympathetic neurotransmitter secretion. The experiments were carried out in isolated superfused field stimulated guinea-pig vas deferens, in which the noradrenaline stores had been labelled by preincubation with tritiated (-)-noradrenaline. Desipramine and normetanephrine were added to prevent rebinding of transmitter. Exogenous noradrenaline was found to cause a dose-dependent and reversible depression of the secretion of tracer noradrenaline evoked by field stimulation. Since the depressing effect was not affected by a ten-fold rise in the desipramine concentration, it seems likely that it was not due to uptake and preferential secretion of unlabelled exogenous noradrenaline, but was tryly due to depression of the secretory mechanism. Adrenaline was significantly more potent than noradrenaline, as inhibitor of the secretion of tracer transmitter, while dopamine, at the same molar concentration, was without effect. The beta-agonist isoprenaline did not depress, but rather tended to enhance, the secretion of tracer noradrenaline. It is concluded that the receptors controlling the secretion of noradrenaline from the sympathetic nerves of guinea-pig vas deferens quantitatively—with regard to sensitivity—as well as qualitatively—with regard to order of preference for different catecholamines—resemble the ‘classical’ alphareceptors of e.g. smooth muscle in the same tissue.