In this study we tested the hypothesis that extracellular ATP regulates the function of the pericyte-containing retinal microvessels. Pericytes, which are more numerous in the retina than in any other tissue, are abluminally located cells that may adjust capillary perfusion by contracting and relaxing. At present, knowledge of the vasoactive molecules that regulate pericyte function is limited. Here, we focused on the actions of extracellular ATP because this nucleotide is a putative glial-to-vascular signal, as well as being a substance released by activated platelets and injured cells. In microvessels freshly isolated from the adult rat retina, we monitored ionic currents via perforated-patch pipettes, measured intracellular calcium levels with the use of fura-2, and visualized microvascular contractions with the aid of time-lapse photography. We found that ATP induced depolarizing changes in the ionic currents, increased calcium levels and caused pericytes to contract. P2X7 receptors and UTP-activated receptors mediated these effects. Consistent with ATP serving as a vasoconstrictor for the pericyte-containing microvasculature of the retina, the microvascular lumen narrowed when an adjacent pericyte contracted. In addition, the sustained activation of P2X7 receptors inhibited cell-to-cell electrotonic transmission within the microvascular networks. Thus, ATP not only affects the contractility of individual pericytes, but also appears to regulate the spatial and temporal dynamics of the vasomotor response.

Pericytes, which are contractile cells located on the outer wall of microvessels, are thought to be particularly important in the retina where the ratio of these cells to vascular endothelial cells is the highest of any tissue. Retinal pericytes are of interest since they may regulate capillary blood flow and because their selective loss is an early event in diabetic retinopathy, which is a common sight‐threatening disorder associated with dysfunction of the blood‐retinal barrier. Although a breakdown in the vascular endothelial barrier is a frequent pathophysiological event, knowledge of the effects of blood‐derived molecules on pericyte function is limited. Based on the premise that ion channels play a vital role in cellular function, we examined the effect of serum on the ionic currents of retinal pericytes. To do this, we used the perforated‐patch configuration of the patch‐clamp technique to monitor the whole‐cell currents of pericytes located on freshly isolated rat retinal microvessels. Exposure to serum reversibly activated inward and outward currents in virtually all of the sampled retinal pericytes. Two types of sustained conductances were induced by serum. These were a calcium‐permeable non‐specific cation (NSC) current and a voltage‐dependent potassium current. In addition, exposure to serum increased the activity of chloride channels which caused transient depolarizing currents. Associated with the activation of these conductances, the membrane potential showed a sustained decrease of 10 ± 2 mV from −56 mV to −46 mV and, also, transient depolarizations to near −30 mV. The serum‐induced depolarizations can activate the voltage‐gated calcium channels expressed by the retinal pericytes. Calcium‐permeable NSC channels appear to play a critical role in the response of pericytes to serum‐derived molecules. Consistent with this, activation of the chloride and potassium channels was sensitive to SK&F 96365, which is a blocker of NSC channels. In addition, chloride and potassium channel activation was dependent on extracellular calcium. The effects of serum on the activity of channels in retinal pericytes were qualitatively mimicked by insulin‐like growth factor‐1 (IGF‐1), which is a normal constituent of the blood. There are significant differences in the effects of serum on retinal pericytes compared with vascular smooth muscle cells. Serum activated sustained conductances in retinal pericytes but not in the vascular smooth muscle cells. This suggests a fundamental difference in the mechanisms by which serum‐derived molecules affect these two types of cells. We conclude that serum‐derived molecules, such as IGF‐1, can activate several types of ion channels in retinal pericytes. These changes in channel activity are likely to influence pericyte function at sites of a breakdown in the blood‐retinal barrier.

We tested the hypothesis that extracellular lactate regulates the function of pericyte-containing retinal microvessels. Although abluminally positioned pericytes appear to adjust capillary perfusion by contracting and relaxing, knowledge of the molecular signals that regulate the contractility of these mural cells is limited. Here, we focused on lactate because this metabolic product is in the retinal extracellular space under both physiological and pathophysiological conditions. In microvessels freshly isolated from the adult rat retina, we used perforated-patch pipettes to monitor ionic currents, fura-2 to measure calcium levels, and time-lapse photography to visualize changes in mural cell contractility and lumen diameter. During lactate exposure, pericyte calcium rose; these cells contracted, and lumens constricted. This contractile response appears to involve a cascade of events resulting in the inhibition of Na+/Ca2+ exchangers (NCXs), the decreased of which function causes pericyte calcium to increase and contraction to be triggered. On the basis of our observation that gap junction uncouplers minimized the lactate-induced rise in pericyte calcium, we propose that the NCXs inhibited by lactate are predominately located in the endothelium. Indicative of the importance of endothelial/pericyte gap junctions, uncouplers of these junctions switched the pericyte response to lactate from contraction to relaxation. In addition, we observed that hypoxia, which closes microvascular gap junctions, also switched lactate's effect from vasocontraction to vasorelaxation. Thus the response of pericyte-containing retinal microvessels to extracellular lactate is metabolically modulated. The ability of lactate to serve as a vasoconstrictor when energy supplies are ample and a vasodilator under hypoxic conditions may be an efficient mechanism to link capillary function with local metabolic need.