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...

Intracellular recordings were made in a pontine slice preparation of the rat brain containing the nucleus locus coeruleus (LC). The pressure application of α,β‐methylene ATP (α,β‐meATP) caused reproducible depolarizations which were depressed by suramin (30 μM) and abolished by suramin (100 μM). Pyridoxal‐phosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS; 10, 30 μM) also concentration‐dependently inhibited the α,β‐meATP‐induced depolarization, although with a much slower time‐course than suramin. Almost complete inhibition developed with 30 μM PPADS. Reactive blue 2 (30 μM) did not alter the effect of α,β‐meATP, while reactive blue 2 (100 μM) slightly depressed it. Pressure‐applied (S)‐α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) also depolarized LC neurones. Kynurenic acid (500 μM) depressed and 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX; 50 μM) abolished the response to AMPA. Suramin (100 μM) potentiated the AMPA effect. Pressure‐applied noradrenaline hyperpolarized LC neurones. Suramin (100 μM) did not alter the effect of noradrenaline. Focal electrical stimulation evoked biphasic synaptic potentials consisting of a fast depolarization (p.s.p.) followed by a slow hyperpolarization (i.p.s.p.). A mixture of D(−)‐2‐amino‐5‐phosphonopentanoic acid (AP‐5; 50 μM), CNQX (50 μM) and picrotoxin (100 μM) depressed both the p.s.p. and the i.p.s.p. Under these conditions suramin (100 μM) markedly inhibited the p.s.p., but did not alter the i.p.s.p. In the combined presence of AP‐5 (50 μM), CNQX (50 μM), picrotoxin (100 μM), strychnine (0.1 μM), tropisetron (0.5 μM) and hexamethonium (100 μM), a high concentration of suramin (300 μM) almost abolished the p.s.p. without changing the i.p.s.p. In the presence of kynurenic acid (500 μM) and picrotoxin (100 μM), PPADS (30 μM) depressed the p.s.p. Moreover, the application of suramin (100 μM) to the PPADS (30 μM)‐containing medium failed to cause any further inhibition. Neither PPADS (30 μM) nor suramin (100 μM) altered the i.p.s.p. It was concluded that the cell somata of LC neurones are endowed with excitatory P2‐purinoceptors. ATP may be released either as the sole transmitter from purinergic neurones terminating at the LC or as a co‐transmitter of noradrenaline from recurrent axon collaterals or dendrites of the LC neurones themselves. British Journal of Pharmacology (1997) 122, 423–430; doi:

No abstract

Taurine, a major osmolyte in the brain evokes a long‐lasting enhancement (LLETAU) of synaptic transmission in hippocampal and cortico‐striatal slices. Hippocampal LLETAU was abolished by the GABA uptake blocker nipecotic acid (NPA) but not by the taurine‐uptake inhibitor guanidinoethyl sulphonate (GES). Striatal LLETAU was sensitive to GES but not to NPA. Semiquantitative PCR analysis and immunohistochemistry revealed that taurine transporter expression is significantly higher in the striatum than in the hippocampus. Taurine transporter‐deficient mice displayed very low taurine levels in both structures and a low ability to develop LLETAU in the striatum, but not in the hippocampus. The different mechanisms of taurine‐induced synaptic plasticity may reflect the different vulnerabilities of these brain regions under pathological conditions that are accompanied by osmotic changes such as hepatic encephalopathy.

Ethanol intoxication results partly from actions of ethanol at specific ligand-gated ion channels. One such channel is the GABA A receptor complex, although ethanol's effects on GABA A receptors are variable. For example, we found that hippocampal neurons from selectively bred mice and rats with high hypnotic sensitivity to ethanol have increased GABA A receptor-mediated synaptic responses during acute ethanol treatment compared with mice and rats that display low behavioral sensitivity to ethanol. Here we investigate whether specific protein kinase C (PKC) isozymes modulate hypnotic and GABA A receptor sensitivity to ethanol. We examined acute effects of ethanol on GABA A receptor-mediated inhibitory postsynaptic currents (IPSCs) in mice lacking either PKC␥ (PKC␥ ). GABA A receptor-mediated IPSCs were evoked in CA1 pyramidal neurons by electrical stimulation in stratum pyramidale, and the responses were recorded in voltage-clamp mode using whole-cell patch recording techniques. Ethanol (80 mM) enhanced the IPSC response amplitude and area in PKC␥ ϩ/ϩ mice, but not in the PKC␥ Ϫ/Ϫ mice. In contrast, ethanol markedly potentiated IPSCs in the PKC⑀ Ϫ/Ϫ mice, but not in PKC⑀ ϩ/ϩ littermates. There was a positive correlation between ethanol potentiation of IPSCs and the ethanol-induced loss of righting reflex such that mice with larger ethanol-induced increases in GABA A receptor-mediated IPSCs also had higher hypnotic sensitivity to ethanol. These results suggest that PKC␥ and PKC⑀ signaling pathways reciprocally modulate both ethanol enhancement of GABA A receptor function and hypnotic sensitivity to ethanol.The mechanisms of ethanol intoxication are complex and involve many regions of the brain. Although alcohol was once thought to act nonselectively to modify lipid mobility in neuronal plasma membranes, it is now clear that ethanol interacts at specific neuronal proteins, including some voltageand ligand-gated ion channels (Lovinger, 1997;Mihic, 1999). In general, acute ethanol treatment decreases excitation via suppression of an N-methyl-D-aspartate-activated current and increases inhibition by enhancing GABA A receptor-mediated conductance, although there is large variability in the reported effects of ethanol on these and other receptor-channel complexes (Crews et al., 1996). The GABA A receptor complex is the primary mediator of fast inhibitory neurotransmission in the central nervous system and is an important target of anesthetic compounds (Mihic et al., 1994;Harris, 1999). Ethanol has a considerable range of effects on GABA A receptor-mediated responses. For example, intoxicating concentrations of ethanol enhance GABA A receptor-mediated Cl Ϫ flux in brain synaptosomal or microsac preparations (Allan and Harris, 1986) and in cultured neurons (Mehta and Ticku, 1994). Electrophysiological studies have shown that ethanol increases GABA A receptor function in some brain preparations (