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...

The spatial distributions of various specific lipids in freeze-dried mouse brain sections were monitored using time-of-flight secondary ion mass spectrometry (TOF-SIMS). Mouse brain sections were prepared by cryosectioning, rinsing in 0.15 M NH3HCOO, and freeze-drying, after which the samples were analyzed directly by TOF-SIMS, using Au3+ ions as primary ions. Positive and negative TOF-SIMS spectra of the tissue surface contained peaks from quasimolecular ions of a variety of specific lipids, including cholesterol, sulfatides, phosphatidylinositols, and phosphatidylcholines. Images showing the spatial signal intensity distributions of specific ions were recorded across analysis areas ranging from 100 x 100 microm(2) to 9 x 9 mm(2). The results demonstrate a highly complementary localization of cholesterol and phosphatidylcholine over dimensions from millimeter to micrometer range. Characteristic spatial distributions of several other lipids, including sulfatides and phosphatidylinositols, were observed. Principal component analysis was used to localize regions of the sample surface that show common spectral features. Spectra from different such regions showed large variations in lipid ion signals, indicating large variations in the lipid composition in different regions.

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The hypothesis that adenosine acting on adenosine A1 receptors (A1R) regulates several renal functions and mediates tubuloglomerular feedback (TGF) was examined using A1R knockout mice. We anesthetized knockout, wild-type, and heterozygous mice and measured glomerular filtration rate, TGF response using the stop-flow pressure (P(sf)) technique, and plasma renin concentration. The A1R knockout mice had an increased blood pressure compared with wild-type and heterozygote mice. Glomerular filtration rate was similar in all genotypes. Proximal tubular P(sf) was decreased from 36.7 +/- 1.2 to 25.3 +/- 1.6 mmHg in the A1R+/+ mice and from 38.1 +/- 1.0 to 27.4 +/- 1.1 mmHg in A1R+/- mice in response to an increase in tubular flow rate from 0 to 35 nl/min. This response was abolished in the homozygous A1R-/- mice (from 39.1 +/- 4.1 to 39.2 +/- 4.5 mmHg). Plasma renin activity was significantly greater in the A1R knockout mice [74.2 +/- 14.3 milli-Goldblatt units (mGU)/ml] mice compared with the wild-type and A1R+/- mice (36.3 +/- 8.5 and 34.1 +/- 9.6 mGU/ml), respectively. The results demonstrate that adenosine acting on A1R is required for TGF and modulates renin release.

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