Hyperalgesia, anxiety, and decreased hypoxic neuroprotection in mice lacking the adenosine A₁receptor

Abstract

Caffeine is believed to act by blocking adenosine A1 and A2A receptors (A1R, A2AR), indicating that some A1 receptors are tonically activated. We generated mice with a targeted disruption of the second coding exon of the A1R (A1R ؊/؊ ). These animals bred and gained weight normally and had a normal heart rate, blood pressure, and body temperature. In most behavioral tests they were similar to A1R ؉/؉ mice, but A1R ؊/؊ mice showed signs of increased anxiety. Electrophysiological recordings from hippocampal slices revealed that both adenosine-mediated inhibition and theophylline-mediated augmentation of excitatory glutamatergic neurotransmission were abolished in A1R ؊/؊ mice. In A1R ؉/؊ mice the potency of adenosine was halved, as was the number of A1R. In A 1R؊/؊ mice, the analgesic effect of intrathecal adenosine was lost, and thermal hyperalgesia was observed, but the analgesic effect of morphine was intact. The decrease in neuronal activity upon hypoxia was reduced both in hippocampal slices and in brainstem, and functional recovery after hypoxia was attenuated. Thus A1Rs do not play an essential role during development, and although they significantly influence synaptic activity, they play a nonessential role in normal physiology. However, under pathophysiological conditions, including noxious stimulation and oxygen deficiency, they are important. A denosine acts on four cloned and pharmacologically characterized receptors, A 1 , A 2A , A 2B , and A 3 (1). Adenosine is believed to play a particularly important role in hypoxia and ischemia, and there is evidence that adenosine serves to limit damage secondary to ATP loss (2, 3). However, adenosine may have important actions under more normal physiological circumstances as well. For instance, the effects of caffeine, at concentrations reached during habitual caffeine consumption, are believed to be a consequence of blockade of tonic activity at some A 1 and A 2A receptors (A 1 R and A 2A R) (4). Studies on mice lacking A 2A Rs show that adenosine tonically activates A 2A Rs and that this activation has functional effects, particularly on behavior, blood pressure, and blood platelets (5). A 1 Rs are more widely distributed than A 2A Rs (4, 6), but despite extensive pharmacological studies their physiological and pathophysiological roles remain unclear. Here we show that A 1 Rs mediate physiological as well as pathophysiological effects of endogenous adenosine. In particular, adenosine acts tonically to activate presynaptic and postsynaptic A 1 Rs to depress synaptic transmission and to reduce nociceptive signaling. At elevated levels seen during hypoxia, adenosine acting at A 1 Rs is responsible for the depression of neuronal activity, and in this situation elimination of A 1 Rs results in impaired functional recovery. Materials and MethodsGeneration of A1R Knockout Mice. A major part of the proteincoding sequence of the mouse A 1 R gene (7) corresponding to exon 6 of the human A 1 R gene described by Ren and Stiles (8) was cloned. The targeting construct was b...

Mice lacking the adenosine A1 receptor have normal spatial learning and plasticity in the CA1 region of the hippocampus, but they habituate more slowly

“…We found that elimination of A 1 Rs clearly affected synaptic physiology in hippocampus, confirming earlier results (Johansson et al, 2001;Masino et al, 2002). As noted previously, hippocampal slices from A 1 R −/− mice were insensitive to the application of exogenous adenosine.…”

“…This adenosine application assessed functionally the genotype of each animal and in each case confirmed the genotype determined by PCR analysis. Previous studies demonstrated that adenosine has no effect on synaptic transmission in hippocampal slices from A 1 R −/− animals, has an intermediate effect in slices A 1 R +/− animals (due to the increased EC 50 for adenosine), and has the expected profound inhibitory effect in slices from A 1 R +/+ animals (Johansson et al, 2001;Masino et al, 2002).…”

“…Mice lacking the second encoding exon of the adenosine A 1 R were generated as described previously (Johansson et al, 2001). They were the offspring of A 1 R +/− mice on a 50% C57BL/ 6, 50% 129/OlaHsd background, which in turn were derived from matings of male chimeric A 1 R −/− mice with normal C57BL/6 females.…”

Adenosine dysfunction in astrogliosis: cause for seizure generation?

“…Animals were kept in a 12-hour light/dark cycle with lights on from 7 am to 7 pm in individually ventilated isolator cages and food and water were supplied ad libitum. Four different genotypes of mice were used in this study, all maintained on a C57BL/6 background: (1) wild type; (2) homozygous Adk-tg mice with brain wide overexpression of a conditional Adk-transgene, but lack of the endogenous Adk gene; these animals are characterized by high ADK expression in CA3 neurons and have been characterized previously (Fedele et al, 2005;Pignataro et al, 2007;Wu and Boison, 2007;Yee et al, 2007;Li et al, 2008); (3) fb-Adk-def mice with reduced levels of forebrain ADK were created based on Emx1-Cre driven recombination of the loxP-flanked Adk-transgene in Adk-tg mice (Li et al, 2008); (4) homozygous and heterozygous A 1 receptor knockout mice (Johansson et al, 2001;Fedele et al, 2006).…”

“…To investigate the possibility for spontaneous seizures in Adk-tg mice, 6 naïve animals were equipped with bipolar recording electrodes implanted into the CA3 region. In parallel, untreated homozygous and heterozygous A 1 receptor knockout mice (Johansson et al, 2001), as well as KA injected wild type mice 3 weeks after SE received intra-CA3 recording electrodes (n = 5 per group). One day after electrode implantation each animal was subjected to 8 hours of continuous EEG recordings.…”