Gap junctions (GJs) are aggregates of ten to thousands of individual intercellular channels that permit the cytoplasmic exchange between adjacent cells. They allow the diffusion of ions, small metabolites (e.g., nicotinamide adenine dinucleotide: NAD+, glucose, lactate, and glutamate), and second messengers (e.g., cyclic adenosine monophosphate [cAMP] and inositol trisphosphate [IP3]) between adjacent cells [40,41,42]. These connections support several biological processes in the animal kingdom, such as the propagation of electrical and chemical signals, which are crucial for cell coordination and survival. GJ channels are formed by integral membrane proteins called connexins, whose family has 21 isoforms in humans that are named according to their molecular weight [42,43]. All connexins feature four transmembrane domains, two extracellular loops, each with three highly conserved cysteine residues, an intracellular loop, and the C- and N-terminal ends on the cytoplasmic domain [42]. Six connexin monomers assemble around a central pore to form a hemichannel (HC) or connexons, whereas the apposition of two connexons from adjacent cells creates an intercellular GJ channel [42]. Cells expressing more than one type of connexin could eventually form heteromeric connexons. Likewise, cells expressing different connexons could form heterotypic GJs [42].

HCs act as free channels in the plasma membrane, favoring the exchange of small molecules and ions between the cytosol and the extracellular space [43,44] (Figure 3). Al-though initially they were catalogued as non-selective channels, nowadays, the evidence seems to indicate the opposite [45,46,47,48]. For example, Cx32 HCs show a slight preference for anions, while Cx40 and Cx43 have a high selectivity for cations [49,50,51]. In the same way, selectivity for ATP is 300-fold higher in Cx43 than in Cx32 HCs; for ADP, glutamate, and glutathione, the selectivity is 20-fold higher in Cx43 than in Cx32 HCs, while adenosine is 10-fold more permeable in Cx32 than Cx43 HCs [42,52]. The physiological opening of these channels is critical for the proper function of multiple biological processes, such as neurite outgrowth [53], neutrophil migration [54], and regulation of acid–base homeostasis in the kidney [55], among others. The HC opening is tightly regulated and relies on changes in voltage, intracellular/extracellular Ca²⁺ concentration ([Ca²⁺]_i) [56], pH [57], post-translational modifications (e.g., phosphorylation [58] and S-nitrosylation [59]), and protein interactions [42]. Nevertheless, persistent activation of HCs has been associated with the genesis and progression of diverse pathological stages [60,61,62]. At least three mechanisms have linked the exacerbated opening of HCs with cell dysfunction and damage [63]. Firstly, the uncontrolled entry of Na⁺ and Cl⁻ through HCs may result in osmotic and ionic imbalances coupled to further cell swelling and plasma membrane breakdown [63,64,65]. Second, some HCs are permeable to Ca²⁺ [66,67], which could permit its influx to the cytosol during pathological conditions. The direct or indirect increase in [Ca²⁺]_i mediated by HCs could lead to Ca²⁺ overload and consequent induction of different proteases, phospholipases, and other hydrolytic enzymes, as well as oxidative stress and caspase activation [68,69]. Finally, exacerbated HC opening may trigger the release of high amounts of potentially toxic molecules to neighboring cells, such as glutamate, in the case of the brain [63,70].

Pannexins (Panxs) belong to a family of transmembrane proteins that bear structural homology with innexins, the GJ protein family of invertebrates [71]. Although connexins and pannexins do not share significant sequence homology, both families exhibit similar transmembrane topology: four transmembrane domains with two extracellular loops and intracellular N- and C-terminals. Unlike connexins, which have several extracellular cysteine residues, pannexins have only two cysteines and a highly glycosylated aspartate in the second extracellular loop, which would hinder GJs formation (see below) [72,73] (Figure 3). Three pannexin isoforms have been identified to date: pannexin-1 (Panx1) (~48 kDa), is the most ubiquitous member of its family, broadly expressed in the brain, muscle, and other tissues [74]; pannexin-2 (Panx2) (~45 kDa), predominantly expressed in the brain; and pannexin-3 (Panx3) (~73 kDa), found in skin, bone, cartilage, skeletal muscle, and blood vessels [72,73,75,76]. Pannexin monomers oligomerize around a central pore in heptamers (Panx1, Panx3) or octamers (Panx2) called pannexons, allowing the exchange of ions and small molecules between the cytoplasm and the extracellular milieu [71,72,77]. Panx1 channels have been historically described as non-junctional membrane channels; nevertheless, recent evidence suggests that Panx1 can form GJs channels due to a differential glycosylated state at its extracellular loop [78]. Sequence analysis of Panx1 reveals that the Asp254 residue can be gradually glycosylated, which yields three different Panx1 bands in Western blot analysis: Gly-0 (non-glycosylated), Gly-1 (several mannose glycosylation), and Gly-2 (complex sugar glycosylation) [78]. Mutagenesis studies show that these glycosylations facilitate Panx1 trafficking but are not essential for the arrival of Panx1 to the plasma membrane or the function of a GJ channel. However, the Gly-2 variant of Panx1 is predominant on the surface [72,78].

Panx1 channels are associated with the regulation of vascular tone and flow [79], mucociliary clearance in the airway epithelial [80], phagocytosis of apoptotic cells [81], activation of the inflammasome, and recruitment of immune cells [82,83]. Similar to HCs, the persistent opening of Panx1 channels has been proposed to be harmful to cells, contributing to cell death in episodes of ischemia [84] or facilitating viral infection [85]. Panx1 channels have been extensively studied as crucial pathways for releasing ATP and pivotal players for purinergic receptor-mediated intracellular Ca²⁺ wave propagation [86,87,88]. Indeed, Panx1 channels release ATP at resting membrane potential and under physiological extracellular concentrations of divalent cations (Ca²⁺ and Mg²⁺) [73]. Furthermore, multiple direct mechanisms activate these channels, including mechanical stimulation [89], caspases-dependent cleavage of the C-tail of Panx1 [81], hypotonicity, ischemia [73], or a secondary response to the activation of receptors ionotropic as P2X4, P2X7, and NMDAR, which are permeable to cations, including Na⁺, K⁺, and Ca²⁺ [83,88,90], or metabotropic receptors such as P2Y1, P2Y2, and P2Y6 [90], protease-activated receptor-1 (PAR-1), bradykinin B₂ receptor (BDKRB2) [91], histamine H₁ receptor (HRH1), and α_1D-adrenergic receptors (α_1D-AR) [92].

3. Connexins and Pannexin-Based Channels in the Kidney