1 Vanilloid receptors (VR1) were cloned from human and rat dorsal root ganglion libraries and expressed in Xenopus oocytes or Chinese Hamster Ovary (CHO) cells. 2 Both rat and human VR1 formed ligand gated channels that were activated by capsaicin with similar EC 50 values. Capsaicin had a lower potency on both channels, when measured electrophysiologically in oocytes compared to CHO cells (oocytes: rat=1.90+0.20 mM; human=1.90+0.30 mM: CHO cells: rat=0.20+0.06 mM; human=0.19+0.08 mM). 3 In CHO cell lines co-expressing either rat or human VR1 and the calcium sensitive, luminescent protein, aequorin, the EC 50 values for capsaicin-induced responses were similar in both cell lines (rat=0.35+0.06 mM, human=0.53+0.03 mM). 4 The threshold for activation by acidic solutions was lower for human VR1 channels than that for rat VR1 (EC 50 pH 5.49+0.04 and pH 5.78+0.09, respectively). 5 The threshold for heat activation was identical (428C) for rat and human VR1. 6 PPAHV was an agonist at rat VR1 (EC 50 between 3 and 10 mM) but was virtually inactive at the human VR1 (EC 50 410 mM). 7 Capsazepine and ruthenium red were both more potent at blocking the capsaicin response of human VR1 than rat VR1. 8 Capsazepine blocked the human but not the rat VR1 response to low pH. Capsazepine was also more e ective at inhibiting the noxious heat response of human than of rat VR1.

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The vanilloid receptor 1 (VR1 or TRPV1) ion channel is activated by noxious heat, low pH and by a variety of vanilloid-related compounds. The antagonist, capsazepine is more effective at inhibiting the human TRPV1 response to pH 5.5 than the rat TRPV1 response to this stimulus. Mutation of rat TRPV1 at three positions in the S3 to S4 region, to the corresponding human amino acid residues I514M, V518L, and M547L decreased the IC 50 values for capsazepine inhibition of the pH 5.5 response from >10,000 nM to 924 ؎ 241 nM in [Ca 2؉ ] i assays and increased capsazepine inhibition of the capsaicin response to levels seen for human TRPV1. We have previously noted that phorbol 12-phenylacetate 13-acetate 20-homovanillate (PPAHV) is a strong agonist of rat TRPV1 but not human TRPV1 in [Ca 2؉ ] i assays (1). Mutation of methionine 547 in S4 of rat TRPV1 to leucine, found in human TRPV1 (M547L), reduced the ability of PPAHV to activate TRPV1 by ϳ20-fold. The reciprocal mutation of human TRPV1 (L547M) enabled the human receptor to respond to PPAHV. These mutations did not significantly affect the agonist activity of capsaicin, resiniferatoxin (RTX) or olvanil in [Ca 2؉ ] i assays. Introducing the equivalent mutation into guinea pig TRPV1 (L549M) increased the agonist potency of PPAHV by > 10-fold in the [Ca 2؉ ] i assay and increased the amplitude of the evoked current. The rat M547L mutation reduced the affinity of RTX binding. Thus, amino acids within the S2-S4 region are important sites of agonist and antagonist interaction with TRPV1.The vanilloid compound capsaicin, the hot substance in chili peppers, activates a specific receptor on sensory nerves termed the vanilloid receptor 1 (VR1 or TRPV1), 1 which was cloned and initially characterized by Caterina et al. (2). TRPV1 is activated by a number of other compounds containing vanillyl groups (see Fig. 1) such as the capsaicin analogue olvanil (3), the phorbol compounds resiniferatoxin (RTX), and phorbol 12-phenyl-acetate 13-acetate 20-homovanillate (PPAHV) (4). A number of endogenous lipid molecules, such as anandamide (5) and lipoxygenase products, including some hydroxyeicosatetraenoic acids and leukotriene B 4 (6), which may be produced during inflammation also activate TRPV1. In addition to these agonists, TRPV1 is activated by low pH (Ͻ6.5) and by noxious heat (Ͼ42°C) (7-9)We have shown differences in the pharmacological properties of the mammalian orthologues of TRPV1 (1, 10). Human, rat and guinea pig TRPV1 expressed in mammalian cells can be similarly activated by capsaicin, low pH, and noxious heat (Ͼ42°C) (1, 2, 10). However, the competitive capsaicin antagonist capsazepine (11, 12) and the agonists PPAHV and RTX show species-specific differences (1, 10, 13). Capsazepine is an effective capsaicin antagonist at TRPV1 from rat, human and guinea pig. While capsazepine blocks the responses of human and guinea pig TRPV1 to low pH and heat, we observed that it only weakly inhibits these responses in cells expressing rat TRPV1 in [Ca 2ϩ ] i assays (1, 10). This pha...

The maximum activities of 5' nucleotidase, adenosine kinase and adenosine deaminase have been measured in several areas of rat and human brain. There is no major difference between the activities of nucleotidase and kinase between rat and human brain, but the activity of deaminase is considerably higher in human brain. The activities of all these enzymes are similar in three areas of rat brain and nine areas of human brain, except for hind brain of the human, which has a low activity of adenosine deaminase. This variation may indicate the existence of different steady-state concentrations of adenosine in certain areas of the brain.Subcellular fractionation of different areas of rat brain showed that, whereas adenosine kinase and deaminase activities were located mainly in the soluble fractions, 5' nucleotidase was present in all subcellular fractions (i.e. membrane, synaptosomal, mitochondria1 and soluble). In particular, there was no major localisation within the synaptosomal fraction. Thus it is unlikely that the regulation of the activities of these enzymes is dependent upon changes within a specific compartment (e.g. synaptosomes) in the brain.