Scheme depicting diminished neurovascular coupling during acute caffeine exposure. There are several potential manifestations related to caffeine restriction of ADO interactions with A₁ and A_2A receptors in presynaptic nerve endings. The net effect of that “multi-receptor blockade” on glutamate (and ATP) release during increased synaptic activity depends on the A₁/A_2A configuration. In an A₁ receptor-dominant presynaptic configuration, the presence of caffeine should remove the postulated A₁ receptor-related block of voltage-dependent Ca²⁺ entry (e.g. [71]), thus increasing Ca²⁺-mediated glutamate and ATP release. In an A_2A receptor-dominant presynaptic scenario (probably the least likely), caffeine blockade is likely to attenuate glutamate release during states of increased neuronal activity. In Fig. 2, the heteromeric A₁/A_2A receptor arrangement is represented. Although precise predictions are not possible, caffeine-related blockade of both receptors might be associated with modest or no changes in activity-evoked glutamate and ATP release. Further reductions in glutamate presence in the synaptic cleft may arise from caffeine blockade of astrocytic A_2A receptors – leading to disinhibition of GLT-1 (permitting greater uptake of glutamate from the synaptic cleft) and diminished A_2A receptor-mediated Ca²⁺-dependent glutamate efflux. A net reduction in glutamate levels in the synaptic cleft could restrict the generation of glutamate receptor-mediated post-synaptic activation. Yet, the presence of caffeine will also prevent ADO-linked post-synaptic depression mediated through activation of A₁ and A_2A receptors. This may permit some post-synaptic activation to occur. Moreover, caffeine blockade of astrocytic A_2A and A_2B receptors, along with the diminished contributions from metabotropic purinergic and glutamatergic receptors (arising from reduced extracellular ATP and glutamate levels), a large reduction in the capacity to generate a Ca²⁺ wave might be expected, resulting in less ADO being “presented” to VSM cells. The diminished ADO exposure, combined with caffeine blockade of VSM A receptors, leaves little capacity remaining for ADO-mediated vasodilation. The potentially diminished capacity to effect “remote” increases in extracellular ADO could also interfere with heterosynaptic influences (see Fig. 1 legend); although the lack of relevant information does not permit any further discussion of this matter. The above speculation provides some (although not the only) possible explanations for the finding that acute caffeine administration profoundly attenuates the in vivo dilation of pial arterioles accompanying somatosensory activation in rats [54].

The neurovascular unit (represented by a synaptic, astrocytic, and vascular component) and the role of adenosine (ADO), via its receptors, in the coupling of enhanced neural activity to arteriolar vascular smooth muscle (VSM) relaxation. Synaptic Component. Increased axonal activity is characterized by enhanced Ca²⁺ entry and accumulation in presynaptic nerve terminals, promoting a Ca²⁺-dependent vesicular release of, for example, glutamate (glu) and ATP from the presynaptic terminal into the synaptic cleft. The released ATP can be rapidly converted to adenosine (ADO) via ectonucleotidases. The increased ADO can engage A₁ and A_2A receptors on pre- and post-synaptic membranes, and it can interact with A_2A and A_2B receptors on adjacent astrocytes (see below). Although A₁ receptors may be expressed on astrocytes and blood vessels, in the Fig. 1 model, those sites are not assigned any functional significance (see Table 1). The glutamate released from the presynaptic terminal can effect post-synaptic activation via engaging metabotropic (mGluR) or ionotropic (NMDA and AMPA) receptors on post-synaptic dendrites. Post-synaptic activation of A₁ and A_2A receptors has been associated with repression of glutamate-linked post-synaptic function [22]. This could act as a “brake” on trans-synaptic signaling. The patterns of A₁ and A_2A receptor expression, as well as the neurotransmitters they modulate, vary among brain structures. Based upon information obtained from cerebrocortical synaptosomes (where evidence indicates the presence of both A₁ and A_2A receptor-mediated modulation of glutamate release [32,33]), the model depicted in Fig. 1 (and Fig. 2) could be taken to represent cerebral cortex. The figure depicts the presence of A₁ and A_2A receptors in close association with one another in the presynaptic nerve ending. This “heteromeric” arrangement represents one of several possibilities, including scenarios where the A₁ or A_2A receptor subtype predominates. In the heteromeric arrangement, it has been postulated that the G_s-linked A_2A receptor not only will activate adenylyl cyclase (AC), but also, via a PKA-independent mechanism [32,33], prevent the G_i/o-linked A₁ receptor from inhibiting AC, especially under conditions of increased neuronal activity and ADO availability [35]. One consequence of this will be a PKA-driven increased Ca²⁺ influx at the presynaptic membrane, overcoming A₁ receptor-linked depression of voltage-dependent Ca²⁺ entry [33], thereby potentiating Ca²⁺-dependent glutamate/ATP release and extracellular ADO generation. Astrocytic Component. The released ATP and glutamate can interact with astrocyte metabotropic P_2Y receptors and mGluR's, respectively, leading to mobilization of Ca²⁺ from intracellular storage sites within astrocytes. In addition, the increased presence of ADO, arising from the released ATP, activates A_2A receptors on astrocytes leading to cAMP/PKA-dependent mobilization of intracellular Ca²⁺ from cellular stores. Adenosine interaction with astrocytic A_2A receptors also can contribute to blockade of the astrocytic glutamate import protein, GLT-1; and promote Ca²⁺-dependent enhancement of glutamate efflux. This should result in further elevations in glutamate levels in the synaptic cleft, as well as contributing to the astrocytic “Ca²⁺ wave”. The figure also speculates that a PKA-linked “boost” to the astrocytic Ca²⁺ mobilization may arise from ADO binding to G_s-linked A_2B receptors. The “wave” of Ca²⁺ generated by the combined influences of glutamatergic, purinergic P_2Y, and adenosinergic mechanisms will ultimately promote ATP release from astrocytes, including remote sites. ATP represents an important signaling molecule in astrocytes. It arises from cellular glucose and O₂ metabolism and can diffuse (along with Ca²⁺) from astrocyte to astrocyte through gap junctions. Additionally, ATP represents perhaps the most important molecule involved in inter-astrocytic communication. Thus, Ca²⁺-dependent release of ATP from one astrocyte interacts with P_2Y receptors on adjacent astrocytes, contributing to the spread of the Ca²⁺ wave. Arteriolar Component. The release of ATP in the vicinity of arterioles is likely to result in rapid formation of ADO and interactions with smooth muscle A₂ receptors. There is little doubt that cerebral arterioles are well-endowed with A₂ receptors. Both A₂ subtypes are likely to be present on cerebral resistance vessels; although the literature seems to favor the A_2A receptor, especially in intraparenchymal and pial arterioles [24,50]. This is reflected in the figure. Principally, A₂ activation generates cAMP, which is not only capable of activating PKA, but cGMP-dependent protein kinase (PKG) as well [46,70]. The increased kinase function is associated with phosphorylation and opening of K⁺ channels, leading to smooth muscle cell hyperpolarization (↓V_m). This lowers intracellular Ca²⁺ levels through a reduction in Ca²⁺ influx via voltage-operated Ca²⁺ channels. Elevated PKA/PKG function also is accompanied by a reduction in the Ca²⁺-sensitivity of contractile proteins (e.g., myosin). The combination of reduced VSM Ca²⁺ levels and diminished sensitivity to Ca²⁺ leads to relaxation. See text for further discussion and additional citations.