Thiopentone inhibits beta-adrenergic responses in myocardial tissue

The purpose of this study was to determine the importance of inhibition of beta-adrenergic function in thiopentone-induced myocardial depression. Using an isolated, electrically stimulated rat left atria model, contractile dose-response curves to thiopentone (200 μM, 400 μM, 600 μM, 800 μM) were shifted to the right in preparations treated with 10^{− 3} M dibutyryl cyclic adenosine monophosphate (cAMP) compared with atria stimulated with 10^{− 6} M isoprenaline, demonstrating that inhibition of beta-adrenergic mechanisms by thiopentone is physiologically important. Depression by thiopentone was similar in atria treated with 10^{− 5} M forskolin compared with preparations stimulated with 10^{− 6} M isoprenaline, indicating that thiopentone does not block beta-adrenergic receptors. It is concluded that thiopentone depresses myocardial function by several mechanisms, one of which involves inhibition of the adenyl cyclase cascade. The adenyl cyclase enzyme is a likely site where thiopentone inhibits the system; however, other components of the cascade may also be involved. L’objectif de cette étude consiste à déterminer l’influence de l’inhibition de l’activité β-adrenergique sur la dépression myocardique induite par le thiopentone. A l’aide d’un modèle constitué d’une oreillette gauche de rat stimulée électriquement, la relation dose-effet du thiopentone sur la contractilité (200 μM, 400 μM, 600 μM, 800 μM) se déplace vers la droite dans des préparations traitées avec de l’adénosine monophosphorique cyclique (cAMP) 10^{− 3} M comparativement à des oreillettes stimulées avec de l’isoprénaline 10^{− 6} M, ce qui démontre que l’inhibition β-adrénergique provoquée par le thiopentone est physiologiquement importante. La dépression de l’oreillette provoquée par le thiopentone est identique à celle que produit la forskoline 10^{− 5} M comparativement à celle de l’isoprénaline 10^{− 6} M, ce qui indique que le thiopentone n’inhibe pas les récepteurs β-adrénergiques. Les auteurs concluent que le thiopentone déprime la fonction myocardique par plusieurs mécanismes qui impliquent l’inhibition de la cascade de l’adényl cyclase. L’inhibition du système se produit vraisemblablement au niveau de l’enzyme adényl cyclase; cependant, il est possible que d’autres éléments de la cascade de l’adényl cyclase soient impliqués.     

分化诱导剂对人肝癌细胞亚细胞组分酪氨酸蛋白激酶的早期影响

研究双丁酰环磷酸腺苷（ｄｂｃＡＭＰ）和全反式维甲酸（ＡＴＲＡ）两种分化诱导剂对人肝癌细胞株ＳＭＭＣ－７７２１亚细胞组分中酪氨酸蛋白激酶（ＴＰＫ）的早期（２４ｈ内）效应。方法用超离心等法制备胞液（ｃ）,胞核（ｎ）和膜性（ｍ）ＴＰＫ酶液,以聚谷氨酸∶酪氨酸（ｐｏ１ｙＥ．Ｙ）４∶ｌ及γ－３２Ｐ－ＡＴＰ为底物测定ＴＰＫ活力。结果对照和ｄｂｃＡＭＰ处理１ｈ使ｃＴＰＫ和ｎＴＰＫ升高,以后对照降至原有水平,而ｄｂｃＡＭＰ处理细胞的ｎＴＰＫ在１２～２４ｈ略低于对照细胞,但ｍＴＰＫ在１ｈ反而降低,在３ｈ（对照）或６ｈ（ｄｂｃＡＭＰ）升至高峰,１２ｈ后ｄｂｃＡＭＰ使ｍＴＰＫ活力低于对照。ＡＴＲＡ作用于ＳＭＭＣ－７７２１细胞后,１ｈ可使三类ＴＰＫ活力均升至高峰,１２ｈ后ｃＴＰＫ和ｎＴＰＫ活力低于对照细胞,但ｍＴＰＫ活力在２４ｈ才低于对照。结论ｄｂｃＡＭＰ和ＡＴＲＡ对不同亚细胞组分ＴＰＫ有不同的时相效应。一般说来,早期（１～６ｈ）有升高作用,而在１２～２４ｈ则有一下降作用,呈现双相效应     

Role of Ca2+-activated K+ channels in acetylcholine-induced dilatation of the basilar artery in vivo

1. We tested the hypothesis that activation of large conductance calcium-activated potassium channels is involved in dilator responses of the basilar artery to acetylcholine in vivo. Using a cranial window in anaesthetized rats, we examined responses of the basilar artery to acetylcholine.
2. Topical application of acetylcholine (10⁻⁶ and 10⁻⁵ M) increased diameter of the basilar artery from 238±7 μm to 268±7 and 288±7 μm, respectively (P<0.05 vs. baseline diameter). Iberiotoxin (10⁻⁸ M), an inhibitor of large conductance calcium-activated potassium channels, did not affect baseline diameter of the basilar artery. In the presence of 10⁻⁸ M iberiotoxin, 10⁻⁶ and 10⁻⁵ M acetylcholine increased diameter of the basilar artery from 239±7 μm to 246±7 and 261±7 μm, respectively. Thus, iberiotoxin attenuated acetylcholine-induced dilatation of the basilar artery (P<0.05).
3. Sodium nitroprusside (10⁻⁷ and 10⁻⁶ M) increased diameter of the basilar artery from 242±9 μm to 310±12 and 374±13 μm, respectively (P<0.05 vs. baseline diameter). In the presence of iberiotoxin (10⁻⁸ M), sodium nitroprusside (10⁻⁷ and 10⁻⁶ M) increased diameter of the basilar artery from 243±6 μm to 259±9 and 311±12 μm, respectively. Thus, iberiotoxin attenuated dilator responses of the basilar artery to sodium nitroprusside (P<0.05).
4. Iberiotoxin partly inhibited dilator responses of the basilar artery to forskolin, a direct activator of adenylate cyclase, but did not affect vasodilatation produced by levcromakalim, a potassium channel opener.
5. These results suggest that dilator responses of the basilar artery to acetylcholine and sodium nitroprusside are mediated, in part, by activation of large conductance calcium-activated potassium channels. Because both acetylcholine and sodium nitroprusside have been shown to activate guanylate cyclase via nitric oxide, activation of large conductance calcium-activated potassium channels may be one of the major mechanisms by which cyclic GMP causes dilatation of the basilar artery in vivo.

     

Interleukin-1β effects on cyclic GMP and cyclic AMP in cultured rat islets of Langerhans — arginine — dependence and relationship to insulin secretion

Summary When islets were cultured with interleukin-1 (1 or 100 pmol/l) for 12 h in arginine-containing medium, cyclic GMP levels were increased 1.6- and 4.5-fold respectively. The arginine analogue, N--nitro-l-arginine methyl ester, which blocks nitric oxide formation and partially reverses inhibition of insulin secretion by 100 pmol/l interleukin-1, largely, but not completely, blocked generation of cyclic GMP. Treatment of islets with 100 pmol/l interleukin-1 for 12 h significantly decreased islet cyclic AMP generation in the absence of isobutylmethylxanthine (from 13.1±0.7 to 9.3±0.8 fmol/g islet protein), this fall was arginine-dependent and may have resulted from an effect on a cyclic AMP phosphodiesterase, since it was masked if isobutylmethylxanthine was present. Isobutylmethylxanthine (0.4 mmol/l) reduced the inhibitory potency of interleukin-1 in 15 h slightly but significantly from 80.5 to 59.0%. The morpholinosydnonimine SIN-1, which is a nitric oxide donor, inhibited insulin secretion, raised islet cyclic GMP and lowered cyclic AMP; its effects were similar to those of interleukin-1. However, 6-anilinoquinoline-5,8-quinone, [LY83583 (1–10 mol/l)], inhibited insulin secretion, and significantly decreased cyclic GMP while 8-bromocyclic GMP stimulated insulin secretion. Both low- and high-dose interleukin-1 treatment give a large arginine-dependent and a small, yet significant, arginine-independent increase in cyclic GMP. The inhibitory effect of SIN-1 or interleukin-1 on insulin secretion seems to depend to a small extent on decreased islet cyclic AMP, though sustained increases in nitric oxide or depleted islet GTP may directly affect the secretory process.     

Comparison of cyclic nucleotide and energy metabolism of intact mouse retina in situ and in vitro.

Whole mouse retina in situ contains approximately 1 μm adenosine 3′,5′-monophosphate (cyclic AMP) and about fivefold higher levels of guanosine 3′,5′-monophosphate (cyclic GMP). Light-adapted retinas have only half as much of both cyclic nucleotides as dark-adapted retinas.Both cyclic AMP and cyclic GMP levels are modified substantially when retinas are removed from the eye and incubated in physiologic buffers. Cyclic GMP levels increase in light- and dark-adapted retinas, but a differential concentration is always maintained, and after a few minutes of incubation it is even greater than that in in situ retinas. In contrast, the difference between cyclic AMP levels obvious in in situ light- and dark-adapted retinas is obscured during the initial minutes of incubation by a transient rise of cyclic AMP in light-adapted retinas, but then becomes apparent again when cyclic AMP levels fall after about 15 min of incubation. Cyclic AMP and cyclic GMP levels decrease rapidly when isolated dark-adapted retinas, in vitro, are exposed to light and the decrease is a function of light intensity. In contrast, cyclic GMP but not cyclic AMP levels increase when light-adapted retinas, in vitro, are placed in darkness.Ischemia causes a marked reduction of ATP and P-creatine levels and elevates cyclic AMP levels in light- and dark-adapted retinas, in situ. Ischemia depresses cyclic GMP levels in dark-adapted retinas, in situ, but has no effect in light-adapted retinas. In incubated retinas, oxygen deprivation produced by NaCN also decreases energy reserves; however, its effect on cyclic AMP is qualitatively similar to that in vivo only after 15 min of incubation. NaCN has little or no effect on cyclic GMP levels in isolated retinas.These data indicate that cyclic nucleotide regulation in intact, incubated retina has many similarities to that of retina, in vivo, and suggest that with due precautions retina maintained, in vitro, is a valid and useful experimental model system for biochemical and biophysical investigations.     

Structural and steric requirements for β-phenethylamines as agonists of the noradrenergic cyclic AMP generating system in the rat limbic forebrain

Summary The present studies were undertaken to assess the structural and steric requirements for -phenethylamines as agonists of the noradrenergic cyclic AMP generating system in slices of the rat limbic forebrain. Significant agonist activity of -phenethylamines requires a -3,4-dihydroxyphenethylamine with a -hydroxyl group in the R configuration. Thus, dopamine did not stimulate the system at concentrations up to 10^–3 M. Moreover, -hydroxyphenethylamines without a 3,4-catechol group (octopamine, phenylephrine, p-hydroxynorephedrine, metaraminol and methoxamine)-though exerting -agonist activity in peripheral tissues-lack agonist activity in this particular cyclic AMP generating system. The effects of (R)-norepinephrine and (R)-isoproterenol at maximal concentrations were not additive. The results lend further support to the view that the cyclic AMP generating system in slices of the limbic forebrain is part of a norepinephrine receptor coupled adenylate cyclase system with a subpopulation of receptors that are in nature.     

Effects of dopamine on cyclic AMP concentration in the anterior pituitary glandin vitro

Summary Effects of different concentrations of dopamine on the cyclic AMP concentration in the rat anterior pituitary gland were investigated in vitro. Low concentrations of dopamine (10^–9–10^–8 mol/l) were found to decrease, whereas the high concentration (10^–5 mol/l) increased the cyclic AMP concentration in pituitaries collected from ovariectomized and estradiol-treated females. In contrast, dopamine had no effect on the anterior pituitary cAMP concentration when pituitaries were collected from ovariectomized rats which had not received estrogen replacement. These data show that the action of dopamine on the anterior pituitary cAMP largely depends on the dopamine concentration and the hormonal state of animals.This paper was supported by the Polish Academy of Sciences Grant No. 10.4.2.01.5.5.     

Effects of adenosine 3′, 5′-cyclic monophosphate on thermoregulation in both rats and rabbits

Summary The effects of intraventricular administration of dibutyryl adenosine 3, 5-cyclic monophosphate (db cyclic AMP) on the thermoregulatory responses of unanesthetized rats and rabbits to different ambient temperatures (T_a) were assessed. Administration of db cyclic AMP (10–60 mM) produced dose-dependent hypothermia in both rats and rabbits at T_a 2–22 °C. The hypothermia in response to db cyclic AMP was due to decreased metabolic heat production and cutaneous vasodilatation. There was no change in respiratory evaporative heat loss. In contrast, in the heat (30–32 °C), db cyclic AMP administration produced dose-dependent hyperthermia in these animals. The hyperthermia was due to increased metabolism (due to muscular shivering) and decreased heat losses. The reduction in heat losses was shown by a decrease in both cutaneous circulation and respiratory evaporative heat loss. The data demonstrate that the thermoregulatory responses induced by central administration of db cyclic AMP are T_a-dependent.     

“Real time” measurement of endogenous dopamine release during short trains of pulses in slices of rat neostriatum and nucleus accumbens : role of autoinhibition

Summary Release of endogenous dopamine elicited in slices of rat neostriatum or nucleus accumbens by a single electric pulse or by trains of 4 or 10 pulses was examined using fast cyclic voltammetry.Single electric pulses gave rise to a marked and transient increase in the extracellular concentration of dopamine in the neostriatum (by 0.43 mol/l) and nucleus accumbens (by 0.39 mol/l). The overflow elicited by subsequent pulses delivered at a frequency of 0.2 Hz caused separate but much smaller peaks of dopamine concentration, whereas the overflow elicited by subsequent pulses delivered at 1 Hz caused only a shoulder in the descending limb of the peak due to pulse 1. Four pulses at 5 Hz produced a monophasic response that was higher than the single pulse-evoked peak. Nomifensine 1 mol/l greatly increased and prolonged the evoked overflow of dopamine. In the absence of nomifensine, metoclopramide 0.3 mol/l did not change the response to a single pulse or 4 pulses delivered at 0.2 Hz but increased the response to 4 or 10 pulses at 1 Hz and to 4 pulses at 5 Hz. In the presence of nomifensine, metoclopramide increased the response to a single pulse as well as, to a greater extent, the response to 4 pulses at 0.2 Hz and 4 pulses at 1 Hz. Sulpiride 1 mol/l produced effects similar to those of metoclopramide in the neostriatum in the presence of nomifensine. During trains of pulses at 0.2 or 1 Hz, metoclopramide and sulpiride did not increase (or increased only slightly in the presence of nomifensine) the initial peak that reflected dopamine overflow elicited by pulse 1, but increased greatly the subsequent peaks (0.2 Hz) or the sholder (1 Hz) that reflected the overflow due to the subsequent pulses.The study demonstrates the release of dopamine in the neostriatum and nucleus accumbens with high temporal resolution so that, at least at low frequency, the release elicited by each pulse in a train can be recognized. As previously concluded from experiments with ³H-dopamine, single pulses elicit a large release whereas subsequent pulses delivered at 0.2 to 5 Hz elicit much smaller release. Presynaptic autoinhibition develops immediately after pulse 1 in trains of appropriately spaced pulses. However, it is only partly responsible for the marked fall in release after pulse 1; other, unknown factors contribute to the decline.The experiments were carried out at the Department of Pharmacology, Queen Mary and Westfield College, London, UK, while N.L. was a visiting scientist Send offprint requests to N. Limberger at the above address     

Regional differences in evoked dopamine efflux in brain slices of rat anterior and posterior caudate putamen

Summary Fast cyclic voltammetry using carbon fibre microelectrodes in rat brain slices, was used to investigate regional differences in electrically-evoked dopamine (DA) efflux at 10 different sites in the anterior caudate putamen (aCPu) and 10 sites in the posterior caudate putamen (pCPu). For each site DA overflow was evoked by both single pulse (1P) stimulation and by trains of 25 pulses applied at a frequency of 50 Hz (25P/50 Hz). Peak DA efflux evoked by 1P was about 58% greater in the aCPu (0.19 mol/l DA) than in the pCPu (0.12 mol/l DA), but showed no mediolateral variation in either region. Peak DA efflux evoked by 25P/50 Hz relative to 1P efflux also varied between the two regions; the aCPu contained predominantly low ratio (25P/50 HZ: 1P) sites ranging from 1.47 to 3.71, whereas in the pCPu these ratios were higher, ranging from 2.73 to 9.40, and were particularly high in the dorsomedial region of the pCPu. Efflux detected in low ratio sites of the aCPu showed little dependence on the frequency (10 to 500 Hz), or the number of pulses (5 to 20) in a train. By contrast DA efflux evoked in high ratio sites of the pCPu responded in a pulse and frequency dependent manner, the maximum ratio (approximately 8 times 1P) being at 20P/20 Hz. Interestingly the frequency response relationship obtained in the pCPu resembled the profile observed in the nucleus accumbens (NAc).Voltammetric evidence and experiments with selective reuptake blockers indicated that only DA was measured in our studies and 5-HT did not significantly contribute to the frequency dependent pattern of efflux detected in high ratio sites of the pCPu, where striatal 5-HT concentrations are highest. Experiments with the selective D₂ receptor antagonists metoclopramide or (–)sulpiride revealed that under our experimental conditions, DA efflux in the aCPu was not modulated by DA autoreceptor activation. By contrast, autoreceptor modulation did occur in high ratio sites of the pCPu at stimulations lasting longer than approximately 1000 ms.These observations support the concept that the caudate putamen is heterogeneously organised with respect to the frequency characteristics of evoked DA release. The factors controlling frequency dependent release under these conditions may be a function of A10 innervation, since high ratio release sites occur in areas where the density of such innervation is greatest, for example, the dorsomedial pCPu. This is supported by the observation that high ratio release sites are also found in the NAc, which receives dopaminergic fibres predominantly from an A10 region. However, the involvement of different regionally distributed transmitters acting on presynaptic receptors involved in the regulation of dopamine release, or differences between nerve terminals in striosomes and matrix, cannot be excluded. Send offprint requests to S. J. Trout at the above address