Inhibition Of N-type Calcium Channels Research Articles

A Comprehensive Insight on Pharmacological Properties of Cilnidipine: A Fourth-generation Calcium Channel Blocker

Abstract: Preventing the development of cardiovascular problems is a key objective of antihypertensive drugs. Many of the non-pressure related coronary risk factors for hypertension are thought to be connected to an increase in sympathetic activity. The sympathetic systems have N-type calcium channels at the nerve terminals that control neurotransmitter release. Cilnidipine is a unique fourth-generation calcium channel blocker with blocking action on both L-/N- type calcium channels. Several L-type calcium channel blockers (Nilvadipine, amlodipine, azelnidipine, nifedipine, etc.) have been used to treat hypertensive patients. Cilnidipine is a novel drug that exerts a hypotensive effect through vasodilation action via blocking L-type calcium channels and potent antisympathetic activity via blocking N-type calcium channels. Inhibiting N-type calcium channels might be a new approach to treating cardiovascular disorders. Therefore, it is expected that cilnidipine may respond well to complicated hypertension. The present review aims to describe the management mechanism of hypertension, and other pharmacological and physicochemical properties of cilnidipine. Cilnidipine has various other beneficial effects such as lipid-lowering effect, reduced white coat effect, improves insulin sensitivity in essential hypertensive patients, ameliorates osteoporosis in ovariectomized hypertensive rats, reduced arterial stiffness, reduced the risk of pedal edema, antinociceptive effects, neuroprotective and renal protective effect, probably through inhibition of N-type calcium channels. Cilnidipine distinguishes itself from other calcium channel blockers due to its wide range of beneficial pharmacological effects. In conclusion, cilnidipine may be more advantageous than other dihydropyridines, such as nisoldipine, amlodipine, azelnidipine, and other antihypertensive drugs.

Inhibition of N-type calcium channels by phenoxyaniline and sulfonamide analogues.

Building on previous investigations, structural modifications to the neuronal calcium ion channel blocker MONIRO-1 and related compounds were conducted that included replacement of the amide linker with an aniline and isosteric sulfonamide moiety, and the previously used strategy of substitution of the guanidinium group with less hydrophilic amine functionalities. A comprehensive SAR study revealed a number of phenoxyaniline and sulfonamide compounds that were more potent or had similar potency for the CaV2.2 and CaV3.2 channel compared to MONIRO-1 when evaluated in a FLIPR-based intracellular calcium response assay. Cytotoxicity investigations indicated that the sulfonamide analogues were well tolerated by Cos-7 cells at dosages required to inhibit both calcium ion channels. The sulfonamide derivatives were the most promising CaV2.2 inhibitors developed by us to date due, possessing high stability in plasma, low toxicity (estimated therapeutic index > 10), favourable CNS MPO scores (4.0-4.4) and high potency and selectivity, thereby, making this class of compounds suitable candidates for future in vivo studies.

N-type calcium channel blockers: a new approach towards the treatment of chronic neuropathic pain

Neuropathic pain (NP) remains maltreated for a wide number of patients by the currently available treatments and little research has been done in finding new drugs for treating NP. Ziconotide (PrialtTM) had been developed as the new drug, which belongs to the class of ω-conotoxin MVIIA. It inhibits N-type calcium channels. Ziconotide is under the last phase of the clinical trial, a new non-narcotic drug for the management of NP. Synthetically it has shown the similarities with ω-conotoxin MVIIA, a constituent of poison found in fish hunting snails (Conus magus). Ziconotide acts by selectively blocking neural N-type voltage-sensitized Ca2+ channels (NVSCCs). Certain herbal drugs also have been studied but no clinical result is there and the study is only limited to preclinical data. This review emphasizes the N-type calcium channel inhibitors, and their mechanisms for blocking calcium channels with their remedial prospects for treating chronic NP.

Steroidal Alkaloids with a Potent Analgesic Effect Based on N-type Calcium Channel Inhibition.

Two distinctive alkaloids with 6/6/6/5/6/6 fused rings, in which a previously unidentified linkage of C-12/23 generates a rigid skeleton, resulting in a new subtype of steroidal alkaloid, were isolated from Veratrum grandiflorum. Compounds 1 and 2 showed potent analgesic effects in vivo, superior to the well-known analgesic, pethidine (Dolantin), likely by inhibiting CaV2.2 voltage-gated calcium channels.

Novel charged sodium and calcium channel inhibitor active against neurogenic inflammation.

Voltage-dependent sodium and calcium channels in pain-initiating nociceptor neurons are attractive targets for new analgesics. We made a permanently charged cationic derivative of an N-type calcium channel-inhibitor. Unlike cationic derivatives of local anesthetic sodium channel blockers like QX-314, this cationic compound inhibited N-type calcium channels more effectively with extracellular than intracellular application. Surprisingly, the compound is also a highly effective sodium channel inhibitor when applied extracellularly, producing more potent inhibition than lidocaine or bupivacaine. The charged inhibitor produced potent and long-lasting analgesia in mouse models of incisional wound and inflammatory pain, inhibited release of the neuropeptide calcitonin gene-related peptide (CGRP) from dorsal root ganglion neurons, and reduced inflammation in a mouse model of allergic asthma, which has a strong neurogenic component. The results show that some cationic molecules applied extracellularly can powerfully inhibit both sodium channels and calcium channels, thereby blocking both nociceptor excitability and pro-inflammatory peptide release.

Intrathecal Therapy for Cancer-Related Pain.

Objective. The increasing incidence of cancer survivorship has shifted treatment of cancer-related pain from short-term analgesia to long-term chronic pain management. As a result, alternatives to oral analgesics, such as intrathecal therapy, may be beneficial for patients with cancer-related pain. The authors review the use of intrathecal therapy in the management of cancer-related pain. Methods. The Medline database was searched for English-language articles that included “ziconotide” or “morphine” AND (“cancer” OR “malignant”) AND “intrathecal” in title or abstract. Available abstracts from scientific congresses in the areas of neuromodulation and oncology were also reviewed. Results. Intrathecal therapy provides pain relief with reduced systemic concerns in patients with cancer-related pain. Patients should undergo multidisciplinary evaluation and, in most cases, drug trialing before intrathecal pump implantation. Morphine, an opioid (µ-opioid receptor antagonist), and ziconotide, a nonopioid (selective N-type calcium channel inhibitor), are both approved for intrathecal analgesia; however, tolerance and safety concerns may deter the use of intrathecal morphine. Ziconotide has also shown efficacy for reduction of cancer-related pain; however, proper dosing and titration must be used to prevent adverse events. There is little information available on use of intrathecal therapies specifically in cancer survivors. Conclusions. Treatment of cancer-related pain has shifted toward chronic pain management strategies, especially among cancer survivors. Intrathecal therapy provides an alternate route of administration of chronic pain medications (e.g., morphine and ziconotide) for cancer patients with and without active disease, although additional research is needed to support effectiveness in cancer survivors.

Molecular basis of toxicity of N-type calcium channel inhibitor MVIIA

MVIIA (ziconotide) is a specific inhibitor of N-type calcium channel, Cav2.2. It is derived from Cone snail and currently used for the treatment of severe chronic pains in patients unresponsive to opioid therapy. However, MVIIA produces severe side-effects, including dizziness, nystagmus, somnolence, abnormal gait, and ataxia, that limit its wider application. We previously identified a novel inhibitor of Cav2.2, ω-conopeptide SO-3, which possesses similar structure and analgesic activity to MVIIA's. To investigate the key residues for MVIIA toxicity, MVIIA/SO-3 hybrids and MVIIA variants carrying mutations in its loop 2 were synthesized. The substitution of MVIIA's loop 1 with the loop 1 of SO-3 resulted in significantly reduced Cav2.2 binding activity in vitro; the replacement of MVIIA loop 2 by the loop 2 of SO-3 not only enhanced the peptide/Cav2.2 binding but also decreased its toxicity on goldfish, attenuated mouse tremor symptom, spontaneous locomotor activity, and coordinated locomotion function. Further mutation analysis and molecular calculation revealed that the toxicity of MVIIA mainly arose from Met12 in the loop 2, and this residue inserts into a hydrophobic hole (Ile300, Phe302 and Leu305) located between repeats II and III of Cav2.2. The combinative mutations of the loop 2 of MVIIA or other ω-conopeptides may be used for future development of more effective Cav2.2 inhibitors with lower side effects.

Discovery of a 1-isopropyltetrahydroisoquinoline derivative as an orally active N-type calcium channel blocker for neuropathic pain

N-type calcium channel blockade is a promising therapeutic approach for the treatment of neuropathic pain. Starting from lead compound (S)-1, we focused our optimization efforts on potency for N-type calcium channel inhibition and improvement of CYP inhibition profile. 2-{[(1-Hydroxycyclohexyl)methyl]amino}-(1R)-(1-isopropyl-6-methoxy-3,4-dihydroisoquinolin-2(1H)-yl)ethanone oxalate ((R)-5r) was identified as a novel orally active small-molecule N-type calcium channel inhibitor with reduced CYP inhibition liability. Oral administration of (R)-5r improved mechanical allodynia in a spinal nerve ligation model of neuropathic pain in rats with an ED50 value of 2.5mg/kg.

Inhibition of N-type calcium channels by fluorophenoxyanilide derivatives.

A set of fluorophenoxyanilides, designed to be simplified analogues of previously reported ω-conotoxin GVIA mimetics, were prepared and tested for N-type calcium channel inhibition in a SH-SY5Y neuroblastoma FLIPR assay. N-type or Cav2.2 channel is a validated target for the treatment of refractory chronic pain. Despite being significantly less complex than the originally designed mimetics, up to a seven-fold improvement in activity was observed.

Comparison of the Effects of Zonisamide, Ethosuximide and Pregabalin in the Chronic Constriction Injury Induced Neuropathic Pain in Rats.

Background:Evidence has been generated that various anticonvulsant agents provide relief of several chronic pain syndromes and therefore as an alternative to opioids, nonsteroidal anti-inflammatory, and tricyclic antidepressant drugs in the treatment of neuropathic pain. The results of these studies thus raise the question of whether all anticonvulsant drugs or particular mechanistic classes may be efficacious in the treatment of neuropathic pain syndromes.Aim:The aim was to compare the clinically used anticonvulsant drugs which are differ in their mechanism of action in a chronic pain model, the chronic constriction injury, in order to determine if all anticonvulsants or only particular mechanistic classes of anticonvulsants are analgesic.Materials and Methods:The study included zonisamide, ethosuximide and pregabalin. All compounds were anticonvulsant with diverse mechanism of actions. The peripheral neuropathic pain was induced by chronic constriction injury of the sciatic nerve in male Sprague-Dawley rats. Zonisamide (80 and 40 mg/kg), ethosuximide (300 and 100 mg/kg), pregabalin (50 and 20 mg/kg), and saline was administered intraperitoneally in respective groups in a blinded, randomized manner from postoperative day (POD) 7-13. Paw withdrawal duration to spontaneous pain, chemical allodynia and mechanical hyperalgesia and paw withdrawal latency to mechanical allodynia and thermal hyperalgesia were tested before drug administration on POD7 and after administration on POD 7, 9, 11 and 13.Results:The present study suggests that these drugs could provide an effective alternative in the treatment of neuropathic pain. However, zonisamide and pregabalin appears to have suitable efficacy to treat a wide spectrum of neuropathic pain condition.Conclusion:The present findings suggest that the inhibition of N-type calcium channels or voltage-gated sodium and T-type calcium channels provides better analgesic potential instead of inhibition of T-type calcium channels alone.

Discovery of novel tetrahydroisoquinoline derivatives as orally active N-type calcium channel blockers with high selectivity for hERG potassium channels

N-type calcium channels represent a promising target for the treatment of neuropathic pain. The selective N-type calcium channel blocker ziconotide ameliorates severe chronic pain but has a narrow therapeutic window and requires intrathecal administration. We identified tetrahydroisoquinoline derivative 1a as a novel potent N-type calcium channel blocker. However, this compound also exhibited potent inhibitory activity against hERG channels. Structural optimizations led to identification of (1S)-(1-cyclohexyl-3,4-dihydroisoquinolin-2(1H)-yl)-2-{[(1-hydroxycyclohexyl)methyl]amino}ethanone ((S)-1h), which exhibited high selectivity for hERG channels while retaining potency for N-type calcium channel inhibition. (S)-1h went on to demonstrate in vivo efficacy as an orally available N-type calcium channel blocker in a rat spinal nerve ligation model of neuropathic pain.

Discovery and SAR of a novel series of 2,4,5,6-tetrahydrocyclopenta[c]pyrazoles as N-type calcium channel inhibitors

A novel series of substituted 2,4,5,6-tetrahydrocyclopenta[c]pyrazoles were investigated as N-type calcium channel blockers (Cav2.2 channels), a chronic pain target. One compound was active in vivo in the rat CFA pain model.

Synthesis and SAR of 4-aminocyclopentapyrrolidines as orally active N-type calcium channel inhibitors for inflammatory and neuropathic pain

A novel series of N-type calcium channel inhibitors have been discovered. Optimization of potency and HT-ADME properties provides 4-aminocyclopentapyrrolidines with analgesic efficacy after oral dosing.

γ-Aminobutyric Acid Type B (GABAB) Receptor Expression Is Needed for Inhibition of N-type (Cav2.2) Calcium Channels by Analgesic α-Conotoxins

α-Conotoxins Vc1.1 and RgIA are small peptides isolated from the venom of marine cone snails. They have effective anti-nociceptive actions in rat models of neuropathic pain. Pharmacological studies in rodent dorsal root ganglion (DRG) show their analgesic effect is mediated by inhibition of N-type (Ca(v)2.2) calcium channels via a pathway involving γ-aminobutyric acid type B (GABA(B)) receptor. However, there is no direct demonstration that functional GABA(B) receptors are needed for inhibition of the Ca(v)2.2 channel by analgesic α-conotoxins. This study examined the effect of the GABA(B) agonist baclofen and α-conotoxins Vc1.1 and RgIA on calcium channel currents after transient knockdown of the GABA(B) receptor using RNA interference. Isolated rat DRG neurons were transfected with small interfering RNAs (siRNA) targeting GABA(B) subunits R1 and R2. Efficient knockdown of GABA(B) receptor expression at mRNA and protein levels was confirmed by quantitative real time PCR (qRT-PCR) and immunocytochemical analysis, respectively. Whole-cell patch clamp recordings conducted 2-4 days after transfection showed that inhibition of N-type calcium channels in response to baclofen, Vc1.1 and RgIA was significantly reduced in GABA(B) receptor knockdown DRG neurons. In contrast, neurons transfected with a scrambled nontargeting siRNA were indistinguishable from untransfected neurons. In the HEK 293 cell heterologous expression system, Vc1.1 and RgIA inhibition of Ca(v)2.2 channels needed functional expression of both human GABA(B) receptor subunits. Together, these results confirm that GABA(B) receptors must be activated for the modulation of N-type (Ca(v)2.2) calcium channels by analgesic α-conotoxins Vc1.1 and RgIA.

Facilitation of sympathetic neurotransmission by phosphatidylinositol-4,5-bisphosphate-dependent regulation of KCNQ channels in rat mesenteric arteries

Sympathetic nerves regulate vascular tone by releasing neurotransmitters into the vasculature. We previously demonstrated that bradykinin facilitates sympathetic neurotransmission in rat mesenteric arteries. Although little is known about the intracellular mechanism modulating this neurotransmission, recent cell line experiments have shown that the KCNQ channel, which is inhibited by the depletion of membrane phosphatidylinositol-4,5-bisphosphate (PIP₂), participates in the control of neurotransmission by bradykinin. In the present study, we examined the mechanism regulating neurotransmitter release from rat perivascular sympathetic nerves. Excitatory junction potentials (EJPs) elicited by repetitive nerve stimulation (1 Hz, 11 pulses, 20 μs, 20-50 V), a measure of sympathetic purinergic neurotransmission, were recorded with a conventional microelectrode technique in rat mesenteric arteries. Bradykinin (10⁻⁷ mol l⁻¹) significantly enhanced the amplitude of EJPs (n=22, P<0.05). This enhancing effect was abolished by N-type calcium-channel inhibition with ω-conotoxin GVIA (2 × 10⁻⁹ mol ⁻¹l, n=8). The blockade of phospholipase C with U-73122 (10(-6) mol l⁻¹, n=17) also eliminated the facilitatory effect of bradykinin. In addition, the effects of bradykinin were diminished by the prevention of PIP₂ resynthesis with wortmannin (10⁻⁵ mol l⁻¹ n=7) or KCNQ channel inhibition with XE-991 (10⁻⁵ mol l⁻¹, n=7). On the other hand, depletion of intracellular calcium stores with cyclopiazonic acid (3 × 10⁻⁶ mol l⁻¹, n=6) or the inhibition of protein kinase C with bisindolylmaleimide-I (10⁻⁶ mol l⁻¹, n=9) did not alter the action of bradykinin. These data demonstrate that the hydrolysis of PIP₂ by phospholipase C, which is activated by G(q/11)-coupled receptors, and subsequent KCNQ channel inhibition enhance sympathetic purinergic neurotransmission presumably via the activation of N-type calcium channels in rat mesenteric arteries.

Structure–activity relationships of trimethoxybenzyl piperazine N-type calcium channel inhibitors

We previously reported the small organic N-type calcium channel blocker NP078585 that while efficacious in animal models for pain, exhibited modest L-type calcium channel selectivity and substantial off-target inhibition against the hERG potassium channel. Structure–activity studies to optimize NP078585 preclinical properties resulted in compound 16, which maintained high potency for N-type calcium channel blockade, and possessed excellent selectivity over the hERG (∼120-fold) and L-type (∼3600-fold) channels. Compound 16 shows significant anti-hyperalgesic activity in the spinal nerve ligation model of neuropathic pain and is also efficacious in the rat formalin model of inflammatory pain.

Evaluation of the Inhibitory Effect of Dihydropyridines on N-type Calcium Channel by Virtual Three-dimensional Pharmacophore Modeling

Currently, a new type of calcium channel blockers, which can inhibit not only L-type calcium channels abundantly expressed in vascular smooth muscles, but also N-type calcium channels that abound in the sympathetic nerve endings, have been developed. In this study, analysis on a like-for-like basis of the L- and N-type calcium channel-inhibitory activity of typical dihydropyridine-type calcium-channel blockers (DHPs) was performed. Moreover, to understand the differences of N-type calcium channel inhibition among DHPs, the binding of DHPs to the channel was investigated by means of hypothetical three-dimensional pharmacophore modeling using multiple calculated low-energy conformers of the DHPs. All of the tested compounds, i.e. cilnidipine (CAS 132203-70-4), efonidipine (CAS 111011-76-8), amlodipine (CAS 111470-99-6), benidipine (CAS 85387-35-5), azelnidipine (CAS 123524-52-7) and nifedipine (CAS 21829-25-4), potently inhibited the L-type calcium channel, whereas only cilnidipine inhibited the N-type calcium channel (IC50 value: 51.2 nM). A virtual three-dimensional structure of the N-type calcium channel was generated by using the structure of the peptide omega-conotoxin GVIA, a standard inhibitor of the channel, and cilnidipine was found to fit well into this pharmacophore model. Lipophilic potential maps of omega-conotoxin GVIA and cilnidipine supported this finding. Conformational overlay of cilnidipine and the other DHPs indicated that amlodipine and nifedipine were not compatible with the pharmacophore model because they did not contain an aromatic ring that was functionally equivalent to Tyr13 of omega-conotoxin GVIA. Azelnidipine, benidipine, and efonidipine, which have this type of aromatic ring, were not positively identified due to intrusions into the excluded volume. Estimation of virtual three-dimensional structures of proteins, such as ion channels, by using standard substrates and/or inhibitors may be a useful method to explore the mechanisms of pharmacological and toxicological effects of substrates and/or inhibitors, and to discover new drugs.

Use of Intrathecal Ziconotide for Frequent Severe and Intractable Sickle Cell Disease-Related Pain

Abstract 1062Many patients with sickle cell disease (SCD) have recurrent, severe vaso-occlusive pain crises, with profound consequences for their quality of life. These pain crises are sometimes responsive to medical management of the underlying disease or to palliative opioid or nonopioid therapies, but in other cases, the pain is intractable. In addition, patients with SCD are prone to chronic, debilitating pain from related conditions.We decided to test intrathecal (IT) ziconotide therapy using an implantable pump with the addition of a Personal Therapy Manager (PTM) patient control accessory in patients with SCD and severe, intractable pain. Ziconotide is a direct and selective inhibitor of N-type calcium channels, believed to reduce signaling in spinal pain pathways. In randomized, controlled trials, ziconotide has been shown to reduce severe chronic pain of various etiologies. We identified 2 patients with SCD and a history of prolonged, intractable pain as suitable candidates. After successful IT ziconotide bolus dose trials demonstrated the utility of ziconotide, these patients were implanted with programmable IT infusion pumps with PTM. Ziconotide was titrated to effect over several weeks.Patient 1 was a 29-year-old female with a history of SCD and hemoglobin of 7, MCV of 83, and platelet count of 297. She had been treated at major academic centers and by private practice hematologists with maximal medical therapy including hydroxyurea and folic acid. She was hospitalized about once a month in the previous 12 months for administration of IV opioids with patient-controlled analgesia (PCA). Currently she has evidence of aseptic necrosis of both hips, not improved by intra-articular steroid injections; she is not a candidate for joint replacements in view of her youth. She underwent a 1-mcg fluoroscopically directed IT bolus trial of ziconotide, with a reduction in her average visual analog scale (VAS) pain score of 9/10 to 0/10 for 48 hours, which was an unusual occurrence for her. She was able to sleep without difficulty and had improvement in her bilateral lower extremity, hip, rib, and back pain. A pump was then implanted for continuous IT infusion of ziconotide. She has not required emergency room visits or hospitalization for sickle cell crises since pump implantation. She is currently at 3.75 mcg/d of ziconotide with a PTM dose of 0.350 mcg every 3 hours and is interested in returning to work. She has not experienced any significant adverse events during the ziconotide titration phase.Patient 2 was a 31-year-old female with a history of hemoglobin S and thalassemia trait. She had bilateral avascular necrosis of the shoulders and hips and had been treated with chronic opioid therapy (fentanyl 50 mcg) but was still being hospitalized monthly for pain management. She had been treated at 2 major academic institutions as well as multiple community hospitals. The patient admitted to previous use of marijuana and cocaine in an effort to control her pain, as well as to obtaining pain medications from multiple physicians. She responded to a 7-mcg IT ziconotide trial with pain reduction from 8/10 to 0/10 for several days and then underwent implantation of a programmable pump with PTM. Following pump implantation she had satisfactory analgesia and did not require any emergency room visits for pain at a stable ziconotide dose of 1.5 mcg/d with a PTM dose of 0.150 mcg every 3 hours. This patient was receiving home refills and developed a pump pocket infection, requiring explantation of her IT infusion pump. Currently, she is poorly controlled on long-acting opioids and has requested reimplantation of her pump with ziconotide monotherapy.Here we describe 2 cases of successful pain management in SCD with IT ziconotide. While receiving continuous IT infusion of ziconotide, both patients were able to be titrated off of high-dose, long-acting opioids. In addition, the PTM proved useful for dosage finding during the titration phase as well as for aborting pain flares. We are in the process of studying a larger number of patients to better define the improvements achieved in functional status, analgesia, and healthcare savings with improved pain management using ziconotide in patients with SCD. Disclosures:McDowell:Azur Pharma: Speakers Bureau.

Increased cavernosal relaxation by Phoneutria nigriventer toxin, PnTx2-6, via activation at NO/cGMP signaling

Erectile dysfunction mechanisms in diabetic patients are multifactorial and often lead to resistance to current therapy. Animal toxins have been used as pharmacological tools to study penile erection. Human accidents involving the venom of Phoneutria nigriventer spider are characterized by priapism. We hypothesize that PnTx2-6 potentiates cavernosal relaxation in diabetic mice by increasing cGMP. This effect is nNOS dependent. Cavernosal strips were contracted with phenylephrine (10−5 M) and relaxed by electrical field stimulation (EFS, 20V, 1–32 Hz) in the presence or absence of PnTx2-6 (10−8 M).Cavernosal strips from nNOS and eNOS knocaut (KO) mice, besides nNOS inhibitor (10−5M), were used to evaluate the role of this enzyme in the potentiation effect evoked by PnTx2-6. Tissue cGMP levels were determined after stimulation with PnTx2-6 in presence or absence of L-NAME (10−4M) and ω-conotoxin GVIA (10−6M), an N-type calcium channel inhibitor. Results showed PnTx2-6 enhanced cavernosal relaxation in diabetic mice (65%) and eNOS KO mice, but not in nNOS KO mice. The toxin effect in the cavernosal relaxation was abolished by nNOS inhibitor. cGMP levels are increased by PnTx2-6, however L-NAME abolished this enhancement as well as ω-conotoxin GVIA. We conclude PnTx2-6 facilitates penile relaxation in diabetic mice through a mechanism dependent on nNOS, probably via increasing NO/cGMP production.

Differential effects of Go and Gi2 on seizure threshold

The inhibitory G protein family, Gi/o, play important roles in neuronal excitability and epileptic pathways. The Gi/o family may protect from seizures by inhibiting neurotransmitter release via inhibition of N‐type calcium channels and activation ofhyperpolerizing potassium channels. However, increased activation of the Gi/o‐coupled GABAB receptor produces absence seizures. The Gi/o family member responsible for each of these physiological effects has not been established. We aim to understand the role of Go and Gi2, two Gi/o family members, in altering seizure thresholds. We utilize mouse models with enhanced signaling through Gi2 and Go due to a genomic knock‐in (G184S) within the α subunit that prevents negative regulation by the Regulators of G protein Signaling. Mice with enhanced Gi2 signaling, but not Go, show reduced seizure severity and death after administration of the competitive GABAA antagonist bicuculline. In a kindling model with repeated injections of subthreshold doses of pentylenetetrazol (35mg/kg) Go G184S mice, but not Gi2 G184S mice, show a more rapid onset of tonic‐clonic seizures and/or death. Together these data indicate opposing roles for these inhibitory G proteins in setting the murine seizure threshold. Supported by RO1‐GM39561 (RRN) and 5T32GM008322 (JMK)