Positive allosteric modulators of α-amino-3-hydroxy-5-methyl-isoxazole-propionic acid (AMPA) receptors facilitate synaptic plasticity and can improve various forms of learning and memory. These modulators show promise as therapeutic agents for the treatment of neurological disorders such as schizophrenia, ADHD, and mental depression. Three classes of positive modulator, the benzamides, the thiadiazides, and the biarylsulfonamides differentially occupy a solvent accessible binding pocket at the interface between the two subunits that form the AMPA receptor ligand-binding pocket. Here, we describe the electrophysiological properties of a new chemotype derived from a structure-based drug design strategy (SBDD), which makes similar receptor interactions compared to previously reported classes of modulator. This pyrazole amide derivative, JAMI1001A, with a promising developability profile, efficaciously modulates AMPA receptor deactivation and desensitization of both flip and flop receptor isoforms.

Positive allosteric modulators of α-amino-3-hydroxy-5-methyl-isoxazole-propionic acid (AMPA) ionotropic glutamate receptors facilitate synaptic plasticity and contribute essentially to learning and memory, properties which make AMPA receptors targets for drug discovery and development. One region at which several different classes of positive allosteric modulators bind lies at the dimer interface between the ligand-binding core of the second, membrane-proximal, extracellular domain of AMPA receptors. This solvent-accessible binding pocket has been the target of drug discovery efforts, leading to the recent delineation of five “subsites” which differentially allow access to modulator moieties, and for which distinct modulator affinities and apparent efficacies are attributed. Here we use the voltage-clamp technique in conjunction with rapid drug application to study the effects of mutants lining subsites “A” and “B” of the allosteric modulator pocket to assess affinity and efficacy of allosteric modulation by cyclothiazide, CX614, CMPDA and CMPDB. A novel analysis of the decay of current produced by the onset of desensitization has allowed us to estimate both affinity and efficacy from single concentrations of modulator. Such an approach may be useful for effective high throughput screening of new target compounds.

Contraction of muscle evokes the exercise pressor reflex (EPR), which is expressed partly by increases in heart rate and arterial pressure. Patients with peripheral artery disease (PAD) show an exaggerated EPR, sometimes report pain when walking and are at risk for cardiac arrthymias. Previous research suggested that reactive oxygen species (ROS) mediate the exaggerated EPR associated with PAD. To examine the effects of ROS on the EPR, we infused a superoxide scavenger, tiron, into the superficial epigastric artery of decerebrated rats. In some, we simulated PAD by ligating a femoral artery for 72 h before the experiment. The peak EPR in 'ligated' rats during saline infusion averaged 31 ± 4 mmHg, whereas the peak EPR in these rats during tiron infusion averaged 13 ± 2 mmHg (n = 12; P < 0.001); the attenuating effect of tiron on the EPR was partly reversed when saline was reinfused into the superficial epigastric artery (21 ± 2 mmHg; P < 0.01 vs. tiron). The peak EPR in 'ligated' rats was also attenuated (n = 7; P < 0.01) by infusion of gp91ds-tat, a peptide that blocks the activity of NAD(P)H oxidase. Tiron infusion had no effect on the EPR in rats with patent femoral arteries (n = 9). Western blots showed that the triceps surae muscles of 'ligated' rats expressed more Nox2 and p67phox, which are components of NADPH oxidase, compared to triceps surae muscles of 'freely perfused' rats. Tiron added to muscle homogenates reduced ROS production in vitro. The results of the present study provide further evidence indicating that ROS mediates the exaggeration of EPR in rats with simulated PAD.

The exercise pressor reflex is initiated by the contraction-induced activation of group III and IV muscle afferents. The reflex is manifested by increases in arterial blood pressure and cardiac output, which, in turn, are generated by increases in the sympathetic outflow to the heart and vasculature and decreases in the vagal outflow to the heart. In previous experiments, we used a pharmacological approach to assess the role played by the acid-sensing ion channel 3 (ASIC3) on group III and IV afferents in evoking the exercise pressor reflex. In the present experiments, we used an alternative approach, namely functional knockout (KO) of the ASIC3 gene, to confirm and extend our previous finding that pharmacological blockade of the ASIC3 had only a small impact on the expression of the exercise pressor reflex when the arterial supply to the contracting hindlimb muscles of rats was patent. Using this alternative approach, we compared the magnitude of the exercise pressor reflex evoked in ASIC3 KO rats with that evoked in their wild-type (WT) counterparts. We found both WT and ASIC3 KO rats displayed similar pressor responses to static contraction (WT, n = 10, +12 ± 2 mmHg; KO, n = 9, +11 ± 2 mmHg) and calcaneal tendon stretch (WT, n = 9, +13 ± 2 mmHg; KO, n = 7, +11 ± 2 mmHg). Likewise, both WT and ASIC3 KO displayed similar pressor responses to intra-arterial injection of 12 mM lactic acid (WT, n = 9, +14 ± 3 mmHg; KO, n = 8, +18 ± 5 mmHg), 24 mM lactic acid (WT, n = 9,+24 ± 2 mmHg; KO, n = 8, +20 ± 5 mmHg), capsaicin (WT, n = 9,+27 ± 5 mmHg; KO, n = 10, +29 ± 5 mmHg), and diprotonated phosphate ([Formula: see text]; WT, n = 6,+22 ± 3 mmHg; KO, n = 6, +32 ± 6 mmHg). We conclude that redundant receptors are responsible for evoking the pressor reflexes arising from group III and IV afferents.

Contraction of freely perfused hind limb muscles in decerebrate rats evokes the exercise pressor reflex, resulting in sympathetic activation and increased blood pressure. This reflex is propagated along mechanically sensitive group III and metabolically sensitive group IV afferent nerve fibers. Recent research by our laboratory has focused on the exaggeration of the exercise pressor reflex in decerebrate rats with simulated peripheral artery disease, which was induced by ligating the femoral artery for 72 h before the start of the experiment. Recently, we showed that ligating the femoral artery increased the responses of single fiber group III and IV triceps surae muscle afferents to static contraction. The objective of this study was to determine if electrical stimulation of group III and IV afferents at frequencies approximating those occurring during static contraction was capable of reflexively increasing arterial blood pressure. We directly stimulated muscle afferents in the absence of muscle contraction for both freely perfused and ligated rats. We established 0.25 Hz as the minimal stimulation frequency to observe a sustained blood pressure response. The blood pressure response increased in a graded fashion as both stimulus frequency and motor threshold were increased. Additionally, we observed similar blood pressure responses from both freely perfused and ligated rats, suggesting that spinal and medullary processing of group III and IV afferent input plays no role in augmenting the pressor response to contraction caused by femoral artery ligation.