Drug Potencies on Partially Purified Brain D2 Dopamine Receptors

To examine the sensitivities of partially purified dopamine receptors to various dopaminergic agonists and antagonists, canine brain striatum dopamine receptors were enriched by isoelectric focusing. The digitonin-solubilized receptors were prelabelled with [3H]spiperone and focused for two time periods. After 5 h (incomplete focusing), radioactive peaks were detected at pH 6 and 9-11. Only the pH 6 peak revealed drug sensitivities expected of D2 receptors. Receptor recovery of the pH 6 peak was 79% with purification being sevenfold. After focusing overnight to equilibrium, the pH 6 peak further separated into peaks at pH 4.6 and 6.8. The receptor was identified only in the pH 4.6 fraction. The recovery of receptors in the pH 4.6 peak was low (10%), indicating little enrichment of the receptor. The rank order of binding of neuroleptics and dopamine agonists to the purified material was similar to that of the original preparation of soluble receptors. Dopamine did not bind to the purified pH 4.6 fraction unless the phosphate buffer (used during focusing) was replaced with Tris buffer. The absence of receptors in the pH 6.8 and pH 10 fractions, although both contained prelabeled [3H]spiperone, indicates the importance of testing agonists and antagonists on each fraction at each step in purification.     

Immunocytochemical detection of 28000-MW calcium-binding protein in horizontal cells of the rat retina

Summary Horizontal cells of rat retina were labeled intensely by a specific antibody to cerebellar calcium-binding protein. The amacrine cells stained very weakly. The presence of calcium-binding protein in horizontal cells could be of interest for the understanding of the feedback action of these cells on photoreceptors.Abbreviations used CaBP calcium-binding protein - DAB 3,3-diaminobenzidine - PAP unlabelled antibody peroxidase-antiperoxidase immunocytochemical complex On leave from the Department of Physiology, University of British Columbia, Vancouver, Canada     

Regulation of the loss of frost hardiness in Pinus radiata by photoperiod and temperature

Abstract. In controlled environments, the interactive effects of warm (16: 8°C, day: night) and cool (12: 4°C, day: night) temperatures and long (13.5 h) and short (10 h) photoperiods on the dehardening of seedlings of Pinus radiata D. Don were investigated. In another experiment, the effect of four photoperiods from 9 to 14 h was examined. In a third, dehardening at constant temperatures from 5 to 17°C was followed. There was no evidence for an interaction between photoperiod and temperature. Dehardening was temporarily delayed by photoperiods below about 10 h, but there was no other quantitative effect of photoperiod. At constant temperatures, the rate of dehardening was initially constant but declined as the minimum summer frost hardiness was reached. In the initial phase the rate of dehardening was a linear function of temperature, increasing from 0.05°C day⁻¹ at 8°C to 0.30 °C day⁻¹ at 17°C. Temperature controlled the loss of frost hardiness by regulating the rate of dehardening.     

Cathepsin B and D activity in stimulated peritoneal macrophages

Summary Highly sensitive and specific synthetic substrates were used to quantitate cathepsin B and D activity in peritoneal macrophages in response to stimulation in vivo with mineral oil and thioglycollate. After intraperitoneal instillation of mineral oil the activity of cathepsin B increased significantly (to 15 300 units/mg protein versus 7 340 in saline controls), reaching values approaching those found in alveolar macrophages (18 400 units/mg protein). Significantly greater stimulation of enzyme activity was obtained after intraperitoneal instillation of thioglycollate (23 600 units/mg protein). Cathepsin D activity also increased significantly after both mineral oil and thioglycollate. However, the increase was moderate (from 806 to about 1 200 units/mg protein), remaining still more than six times lower-than in alveolar macrophages. The data are the first to demonstrate that cathepsin B activity can be stimulated in vivo in peritoneal macrophages by instillation of agents that induce acute inflammation. They also point to a differential control of expression of cathepsin B and D activity in both peritoneal and alveolar macrophages in spite of the common lysosomal origin of the two enzymes.Abbreviations Cbz -N-benxyloxycarbonyl - 2NA 2-naphthylamine - EDTA ethylenediamine tetraacetate - DMSO dimethylsulfoxide - PBS phosphate-buffered saline - PM peritoneal macrophage - AM alveolar macrophage     

Open-field behaviour in chickens: A replication revisited

In an attempt to show that the open field can still be used as a valid measure of fear, Jones (1983) has reported a failure to replicate some of our findings. The present studies show that this was due to procedural and methodological differences. For instance, we found that birds tested in a novel environment behaved quite differently from those, as in Jones' case, which were placed in one resembling the home cage. Moreover, birds housed in isolation for two days prior to testing reacted differently than those, as again in Jones' case, which were reared in isolation from hatching to the time of testing. The results were interpreted as being consistent with our view that open-field behaviour reflects a conflict between the need to reinstate contact with conspecifics on the one hand, and evade predation on the other.     

125I] 3-quinuclidinyl 4-iodobenzilate: a high affinity, high specific activity radioligand for the M1 and M2-acetylcholine receptors

We have prepared a radioiodinated ligand which binds with high affinity to the muscarinic acetylcholine receptor (m-AChR). A derivative of 3-quinuclidinyl benzilate, [125I] labeled (R) 1-aza-bicyclo(2.2.2)oct-3-yl (R,S)-alpha-hydroxy-alpha-(4-[125I]iodophenyl)phenyl acetate (4- IQNB ) exhibits an affinity for the m-AChR from corpus striatum higher than that of (R) [3H] QNB. Additionally, [125I] 4- IQNB exhibits receptor selectivity for the M1 receptor since the affinity for the receptor from dog and rat heart is lower than that using dog or rat corpus striatum.     

Naltrexone prevents footshock-induced performance deficit in rats

Three experiments demonstrate that inescapable footshock delivered to unrestrained rats produces analgesia as well as performance deficits in subsequent one-way shuttle acquisition. Both the performance and the antinociceptive effects are prevented by pretreatment with as little as 0.1 mg/kg i.p. of the opiate antagonist, naltrexone. These studies suggest that both effects are mediated through opiate receptors with similar underlying naltrexone pharmacodynamics.     

Biochemical significance of enhanced activity of fluorinated 1,25-dihydroxyvitamin D3 in human cultured cell lines

Several human cancer cells possess receptors for 1,25-dihydroxyvitamin D3[1,25-(OH)2D3]. In these cells 1,25-(OH)2D3 has a biphasic concentration-dependent regulatory effect on cell replication and specifically induces its own metabolism. We have studied the effects on these parameters of the native hormone together with those of two analogues fluorinated at the 24-carbon and of 1,24R,25-trihydroxyvitamin D3[1,24R,25-(OH)3D3]. The difluorinated analogue 24,24-difluoro-1,25-(OH)2D3[24,24-F2-1,25-(OH)2D3] is an approximately fivefold more potent inhibitor of cellular replication than the native hormone, while 1,24R,25-(OH)3D3 is about fivefold less potent. This enhanced potency of the fluorinated analogue parallels its enhanced potency in in vivo studies of its effects on calcium and mineral metabolism. However, although the analogue retains replication stimulatory activity, it is clearly no more potent than the native hormone in this activity: 1,24R,25-(OH)3D3 has no significant stimulatory activity. Exposure of the cells to 1,25-(OH)2D3 at 0.05 nM for 6 h increases the subsequent conversion of labelled hormone to aqueous phase soluble compounds by 6.7-fold. None of the other compounds had a similar effect at this concentration. At 10 nM all 1-hydroxylated compounds increased aqueous phase radioactivity about equally (13 to 17-fold); this effect is still specific since 25-OH D3 had no such effect even at 10 nM. Studies on the effects of the fluorinated analogues upon receptor binding of hormone in cell cytosols and uptake of hormone by intact cells clearly demonstrate that the enhanced activity of these analogues is not due to higher receptor affinity or more rapid access to intracellular receptor.(ABSTRACT TRUNCATED AT 250 WORDS)