Pergamon Press
Life Sciences, Vol . 23, pp . 1929-1934 Printed in the U .S .A .
THE EFFECTS OF CADMIUM ON ADRENERGIC NEUROTRANSMISSION IN VITRO* Betty J . Williams, David J . Laubach, Bohdan R . Nechay, and Odd S . Steinsland Department of Pharmacology and Toxicology, The University of Texas Medical Branch, Galveston, Texas 77550 (Received in final form September 15, 1978) Surtmary The effect of cadmium on the response of isolated perfused rabbit ear arteries to nerve stimulation and norepinephrine administration was examined . Cadmium in concentrations of .075- .25UM caused enhancement of the pressor responses to nerve stimulation, but higher concentrations caused Inhibition of the response . The pressor response to norepinephrine was also inhibited by cadmium, but required a 100x higher concentration than that needed for Inhibition of the response to nerve stimulation . The dual effect of cadmium on the response to nerve stimulation suggests a plausible explanation for the conflicting reports in the literature regarding the blood pressure effects of cadmium exposure . The enhancement by low concentrations of cadmium of the response to nerve stimulation provides a possible mechanism for cadmiuminduced hypertension . Recently, we observed that tail arteries and aortic rings isolated from rats chronically exposed to low doses of cadmium exhibited an increased sensitivity to adrenergic nerve stimulation and norepinephrine as compared to arteries and aortic rings isolated from rats that received no cadmium (1) . Isolated tissue studies by other investigators have shown that relatively high concentrations of cadmium (>100UM) inhibit the constrictor response of aortic strips to norepinephrine, epinephrine, angiotensin, and other vasopressor agents (2-4) . At lower concentrations (5-20UM), cadmium has been shown to reduce the pressor response of isolated, rabbit saphenous arteries to adrenergic nerve stimulation (5) ; this effect has been attributed to the inhibition of neurotransmitter release . The present study was undertaken to investigate the acute effects of cadmium, especially at very low concentrations, on adrenergic neurotransmission in isolated, perfused arteries . Methods New Zealand white rabbits (male, 2-3 kg) were sacrificed by a blow to the head . The proximal portion (2-3 cm) of the central ear artery was dissected free, cannulated at both ends, and mounted in a perfusion chamber designed to allow continuous and simultaneous intraluminal perfusion and The preparation and perfusion chamber have been extraluminal "superfusion" . described previously (6) . The preparation was continuously bathed in a *Presented In part at the Fall Meeting of the Southwestern Association of Toxicologists, November 12, 1977 . 0300-9653/78/1106-1929502 .00/0 Copyright (c) 1978 Pergamon Press
1930
Cadmium on Adrenergic Neurotransmission
Vol . 23, No . 19, 1978
perfusion fluid of the following millimolar composition : NaCI, 119 ; NaHC0 3 ~ 25 ; KCI, 4 .7 ; MgS04, 1 .5 ; KH2P04, 1,2 ; CaC12, 2,5 ; glucose, 11 ; disodlum ethylenediaminetetraacetate, 0 .03 : sodium ascorbate, 0 .005 . The perfusion fluid was equilibrated with a 95$ Op-5$ C02 gas mixture . Stimulation of the adrenergic nerve endings was accomplished with rectangular pulses of supramaximal voltage (60-70v) and 1 msec duration from a Grass S88 stimulator . Since the nerve endings are located near the medial advential border, cadmium was administered extraluminally in experiments on the effect of cadmium on the response to nerve stimulation . Direct stimulation of the smooth muscle with norepinephrine was accomplished by brief, intermittent perfusion of norepinephrine intraluminally . Cadmium was administered intraluminally in experiments using norepinephrine ; extraluminal cadmium administration had the same qualitative effect . Solutions containing cadmium, as CdC12, or 1-noreplnephrine bitartrate were prepared fresh daily . The responses during cadmium administration were compared to a series of stable, reproducible control responses obtained immediately prior to cadmium administration . The responses during cadmium administration were expressed as a percent of the control response . Standard errors were calculated for all mean values reported . Data were analyzed using the student's "t" test . Results Addition of cadmium to isolated ear arteries caused both an enhancement and inhibition of the response to nerve stimulation ; the character of the effect was dependent on cadmium concentration . Figure 1 is a tracing from a representative experiment showing the effect of different concentrations of cadmium on the pressor response of the rabbit ear artery to brief, intermittent field stimulation of the sympathetic nerve endings . Each concentration of cadmium was administered during three to four successive stimulations (12-16 min .) . As shown, cadmium at very low concentrations (0 .075-0 .75uM) enhanced the response of the artery to nerve stimulation, but at concentrations above 0 .75vM caused a concentration dependent inhibition of the pressor response . The response to nerve stimulation was almost completely abolished by cadmium at a concentration of 25uM . After cadmium administration was discontinued, the response to nerve stimulation gradually returned, and within twenty minutes had reached the control level . After sixty minutes of continuous perfusion with cadmium free solution the response to nerve stimulation had increased to 140$ of the control . This enhancement persisted for the duration of the experiment . Subsequent administrations of low concentrations of cadmium to this preparation had either no effect or only an inhibitory effect on the response to nerve stimulation . In ten experiments of the type illustrated in figure l, very low concentrations of cadmium (0 .025-0 .75uM) caused a mean increase of 6 .1±2 .1$ (P
Life Sciences, Vol . 23, pp . 1929-1934 Printed in the U .S .A .
THE EFFECTS OF CADMIUM ON ADRENERGIC NEUROTRANSMISSION IN VITRO* Betty J . Williams, David J . Laubach, Bohdan R . Nechay, and Odd S . Steinsland Department of Pharmacology and Toxicology, The University of Texas Medical Branch, Galveston, Texas 77550 (Received in final form September 15, 1978) Surtmary The effect of cadmium on the response of isolated perfused rabbit ear arteries to nerve stimulation and norepinephrine administration was examined . Cadmium in concentrations of .075- .25UM caused enhancement of the pressor responses to nerve stimulation, but higher concentrations caused Inhibition of the response . The pressor response to norepinephrine was also inhibited by cadmium, but required a 100x higher concentration than that needed for Inhibition of the response to nerve stimulation . The dual effect of cadmium on the response to nerve stimulation suggests a plausible explanation for the conflicting reports in the literature regarding the blood pressure effects of cadmium exposure . The enhancement by low concentrations of cadmium of the response to nerve stimulation provides a possible mechanism for cadmiuminduced hypertension . Recently, we observed that tail arteries and aortic rings isolated from rats chronically exposed to low doses of cadmium exhibited an increased sensitivity to adrenergic nerve stimulation and norepinephrine as compared to arteries and aortic rings isolated from rats that received no cadmium (1) . Isolated tissue studies by other investigators have shown that relatively high concentrations of cadmium (>100UM) inhibit the constrictor response of aortic strips to norepinephrine, epinephrine, angiotensin, and other vasopressor agents (2-4) . At lower concentrations (5-20UM), cadmium has been shown to reduce the pressor response of isolated, rabbit saphenous arteries to adrenergic nerve stimulation (5) ; this effect has been attributed to the inhibition of neurotransmitter release . The present study was undertaken to investigate the acute effects of cadmium, especially at very low concentrations, on adrenergic neurotransmission in isolated, perfused arteries . Methods New Zealand white rabbits (male, 2-3 kg) were sacrificed by a blow to the head . The proximal portion (2-3 cm) of the central ear artery was dissected free, cannulated at both ends, and mounted in a perfusion chamber designed to allow continuous and simultaneous intraluminal perfusion and The preparation and perfusion chamber have been extraluminal "superfusion" . described previously (6) . The preparation was continuously bathed in a *Presented In part at the Fall Meeting of the Southwestern Association of Toxicologists, November 12, 1977 . 0300-9653/78/1106-1929502 .00/0 Copyright (c) 1978 Pergamon Press
1930
Cadmium on Adrenergic Neurotransmission
Vol . 23, No . 19, 1978
perfusion fluid of the following millimolar composition : NaCI, 119 ; NaHC0 3 ~ 25 ; KCI, 4 .7 ; MgS04, 1 .5 ; KH2P04, 1,2 ; CaC12, 2,5 ; glucose, 11 ; disodlum ethylenediaminetetraacetate, 0 .03 : sodium ascorbate, 0 .005 . The perfusion fluid was equilibrated with a 95$ Op-5$ C02 gas mixture . Stimulation of the adrenergic nerve endings was accomplished with rectangular pulses of supramaximal voltage (60-70v) and 1 msec duration from a Grass S88 stimulator . Since the nerve endings are located near the medial advential border, cadmium was administered extraluminally in experiments on the effect of cadmium on the response to nerve stimulation . Direct stimulation of the smooth muscle with norepinephrine was accomplished by brief, intermittent perfusion of norepinephrine intraluminally . Cadmium was administered intraluminally in experiments using norepinephrine ; extraluminal cadmium administration had the same qualitative effect . Solutions containing cadmium, as CdC12, or 1-noreplnephrine bitartrate were prepared fresh daily . The responses during cadmium administration were compared to a series of stable, reproducible control responses obtained immediately prior to cadmium administration . The responses during cadmium administration were expressed as a percent of the control response . Standard errors were calculated for all mean values reported . Data were analyzed using the student's "t" test . Results Addition of cadmium to isolated ear arteries caused both an enhancement and inhibition of the response to nerve stimulation ; the character of the effect was dependent on cadmium concentration . Figure 1 is a tracing from a representative experiment showing the effect of different concentrations of cadmium on the pressor response of the rabbit ear artery to brief, intermittent field stimulation of the sympathetic nerve endings . Each concentration of cadmium was administered during three to four successive stimulations (12-16 min .) . As shown, cadmium at very low concentrations (0 .075-0 .75uM) enhanced the response of the artery to nerve stimulation, but at concentrations above 0 .75vM caused a concentration dependent inhibition of the pressor response . The response to nerve stimulation was almost completely abolished by cadmium at a concentration of 25uM . After cadmium administration was discontinued, the response to nerve stimulation gradually returned, and within twenty minutes had reached the control level . After sixty minutes of continuous perfusion with cadmium free solution the response to nerve stimulation had increased to 140$ of the control . This enhancement persisted for the duration of the experiment . Subsequent administrations of low concentrations of cadmium to this preparation had either no effect or only an inhibitory effect on the response to nerve stimulation . In ten experiments of the type illustrated in figure l, very low concentrations of cadmium (0 .025-0 .75uM) caused a mean increase of 6 .1±2 .1$ (P
