Astrocytic β2-adrenergic receptors: From physiology to pathology

Evidence accumulates for a key role of the β₂-adrenergic receptors in the many homeostatic and neuroprotective functions of astrocytes, including glycogen metabolism, regulation of immune responses, release of neurotrophic factors, and the astrogliosis that occurs in response to neuronal injury. A dysregulation of the astrocytic β₂-adrenergic-pathway is suspected to contribute to the physiopathology of a number of prevalent and devastating neurological conditions such as multiple sclerosis, Alzheimer's disease, human immunodeficiency virus encephalitis, stroke and hepatic encephalopathy. In this review we focus on the physiological functions of astrocytic β₂-adrenergic receptors, and their possible impact in disease states.     

Antagonism of α1-adrenergic and serotonergic receptors in the hypoglossal motor nucleus does not prevent motoneuronal activation elicited from the posterior hypothalamus

The perifornical (PF) region of the posterior hypothalamus plays an important role in the regulation of sleep–wake states and motor activity. Disinhibition of PF neurons by the GABA_A receptor antagonist, bicuculline, has been used to study the mechanisms of wake- and motor activity-promoting effects that emanate from the PF region. Bicuculline activates PF neurons, including the orexin-containing cells that have major excitatory projections to brainstem noradrenergic and serotonergic neurons. Since premotor aminergic neurons are an important source of motoneuronal activation, we hypothesized that they mediate the excitation of motoneurons that results from disinhibition of PF neurons with bicuculline. In urethane-anesthetized, paralyzed and artificially ventilated rats, we found that PF bicuculline injections (1 mM, 20 nl) made after combined microinjections into the hypoglossal (XII) nucleus of α₁-adrenergic and serotonergic receptor antagonists (prazosin and methysergide) increased XII nerve activity by 80 ± 16% (SE) of the control activity level. Thus, activation of XII motoneurons originating in the hypothalamic PF region was not abolished despite effective elimination by the aminergic antagonists of the endogenous noradrenergic and serotonergic excitatory drives to XII motoneurons and abolition of XII motoneuronal activation by exogenous serotonin or phenylephrine. These results show that a major component of XII motoneuronal activation originating in the posterior hypothalamus is mediated by pathways other than the noradrenergic and serotonergic projections to motoneurons.     

Heat loss responses and blockade of prostaglandin E2-induced thermogenesis elicited by α1-adrenergic activation in the rostromedial preoptic area

The unilateral microinjection of noradrenaline (NA), but not vehicle solution, into the rostromedial preoptic area (POA) elicited simultaneous increases in cutaneous temperatures of the tail and sole of the foot and decreases in the whole-body O₂ consumption rate, heart rate, and colonic temperature in urethane–chloralose-anesthetized rats, suggesting a coordinate increase in heat loss and decrease in heat production. The magnitude of these responses increased dose-dependently over the range of 1–100 pmol, except for the metabolic and bradycardic responses. Similar hypothermic responses were elicited by the microinjection of 40 pmol methoxamine (an α₁-adrenergic agonist), but not by that of clonidine (an α₂-agonist) or isoproterenol (a β-agonist). Sites at which microinjection of NA elicited hypothermic responses were in the vicinity of the organum vasculosum of the lamina terminalis including the median preoptic nucleus, whereas no thermal or metabolic response was elicited when NA was microinjected into the lateral POA or caudal part of the medial POA. The microinjection of 130 fmol prostaglandin (PG) E₂ into the NA-sensitive site always elicited thermogenic, tachycardic, and hyperthermic responses. Furthermore, the PGE₂-induced febrile responses were greatly attenuated by prior administration of NA at the same site. These results demonstrate that NA in the rostromedial POA exerts α₁-adrenoceptor-mediated hypothermic effects and opposes PGE₂-induced fever.     

Testosterone is essential for α2-adrenoceptor-induced antinociception in the trigeminal region of the male rat

Activation of the α₂-adrenoceptor has been shown to produce antinociception. We have previously shown that the antinociceptive effect of clonidine, an α₂-adrenoceptor agonist, is sex-specific and is abolished by exogenous estrogen in ovariectomized rats or high level of endogenous estrogen in proestrous females. Here, we investigated whether testosterone mediates the antinociceptive effect of clonidine in the trigeminal region of the male rat. Clonidine (7 μg/5 μl) was injected intracisternally through a PE-10 cannula implanted dorsal to the trigeminal region in orchidectomized (GDX) male Sprague–Dawley rats. In separate groups, testosterone propionate (250 μg/100 μl; GDX+T) or β-estradiol benzoate (100 μg/100 μl; GDX+E) were injected subcutaneously 24 and 48 h respectively prior to the N-methyl-d-aspartic acid (NMDA)—or heat-evoked nociceptive test. NMDA-induced number of scratches or duration of scratching behavior did not change significantly in control groups with or without hormonal replacement. Clonidine significantly reduced both measures only in the GDX+T group but not in GDX or GDX+E group. Clonidine also significantly increased head withdrawal latency (HWL) in the GDX+T group, but not in GDX or GDX+E group. The antinociceptive effect of clonidine was reversed by yohimbine, an α₂-adrenoceptor antagonist, in GDX+T group. We conclude that testosterone is required for the expression of antinociception produced by selective activation of the α₂-adrenoceptor in the trigeminal region of the male rat. These findings further our understanding of sex-related differences in the modulation of nociception and may provide insight into development and administration of analgesic agents in young vs. aging men.     

Modulation of potassium channels via the α1B-adrenergic receptor in human osteoblasts

Recent studies have demonstrated the involvement of the sympathetic nerve system in bone metabolism. We have previously demonstrated the expression of adrenergic receptors in the human osteoblast SaM-1 cell line. The aim of this study was to reveal the function of these receptors in osteoblasts using electrophysiological methods. During whole-cell patch clamp recording, the application of noradrenaline reduced the currents induced by a voltage ramp, and the effect was larger in the positive potential range. Pretreatment with the selective α_1B-adrenergic receptor antagonist chloroethylclonidine eliminated the inhibitory effect of noradrenaline. Using Cs-based pipette solution to block potassium channels, the inhibitory effect of noradrenaline disappeared. In contrast, the effect was unaffected in the presence of tetraethylammonium (TEA), a potassium channel blocker. These results suggest that noradrenaline suppresses Cs-sensitive and TEA-insensitive potassium channels via the α_1B-adrenergic receptor in human osteoblasts.     

Amplified respiratory–sympathetic coupling in the spontaneously hypertensive rat: does it contribute to hypertension?

Sympathetic nerve activity (SNA) is elevated in established hypertension. We tested the hypothesis that SNA is elevated in neonate and juvenile spontaneously hypertensive (SH) rats prior to the development of hypertension, and that this may be due to augmented respiratory–sympathetic coupling. Using the working heart–brainstem preparation, perfusion pressure, phrenic nerve activity and thoracic (T8) SNA were recorded in male SH rats and normotensive Wistar–Kyoto (WKY) rats at three ages: neonates (postnatal day 9–16), 3 weeks old and 5 weeks old. Perfusion pressure was higher in SH rats at all ages reflecting higher vascular resistance. The amplitude of respiratory-related bursts of SNA was greater in SH rats at all ages ( P < 0.05). This was reflected in larger Traube–Hering pressure waves in SH rats (1.4 ± 0.8 versus 9.8 ± 1.5 mmHg WKY versus SH rat, 5 weeks old, n = 5 per group, P < 0.01). Recovery from hypocapnic-induced apnoea and reinstatement of Traube–Hering waves produced a significantly greater increase in perfusion pressure in SH rats ( P < 0.05). Differences in respiratory–sympathetic coupling in the SH rat were not secondary to changes in central or peripheral chemoreflex sensitivity, nor were they related to altered arterial baroreflex function. We have shown that increased SNA is already present in SH rats in early postnatal life as revealed by augmented respiratory modulation of SNA. This is reflected in an increased magnitude of Traube–Hering waves resulting in elevated perfusion pressure in the SH rat. We suggest that the amplified respiratory-related bursts of SNA seen in the neonate and juvenile SH rat may be causal in the development of their hypertension.     

Activation of α2-adrenoceptors in the lateral hypothalamus reduces pilocarpine-induced salivation in rats

Anti-hypertensive drugs that act on central α₂-adrenoceptors and imidazoline receptors usually cause dry mouth in patients. A central area important for the control of salivary secretion and also for the effects of α₂-adrenoceptor activation is the lateral hypothalamus (LH). Therefore, in the present study we investigated the effects of the injections of moxonidine (an α₂-adrenoceptor and imidazoline agonist) alone or combined with RX 821002 (α₂-adrenoceptor antagonist) into the LH on the salivation induced by intraperitoneal (i.p.) pilocarpine (cholinergic muscarinic agonist). Male Holtzman rats with stainless steel cannula implanted into the LH were used. Saliva was collected using pre-weighted small cotton balls inserted into the animal’s mouth under ketamine anesthesia. Salivation induced by i.p. pilorcarpine (4 μmol/kg of body weight) was reduced by the injection of moxonidine (10 and 20 nmol/0.5 μl) into the LH (222 ± 46 and 183 ± 19 mg/7 min, vs. vehicle: 480 ± 30 mg/7 min). The inhibitory effect of moxonidine on pilocarpine-induced salivation was abolished by prior injections of RX 821002 (160 and 320 nmol/0.5 μl) into the LH (357 ± 25 and 446 ± 38 mg/7 min). Injections of the α₁-adrenoceptor antagonist prazosin (320 nmol/0.5 μl) into the LH did not change the effects of moxonidine. The results show that activation of α₂-adrenoceptors in the LH inhibits pilocarpine-induced salivation, suggesting that LH is one of the possible central sites involved in the anti-salivatory effects produced by the treatment with α₂-adrenoceptor agonists.     

Role of α2/δ subunit in the development of morphine-induced rewarding effect and behavioral sensitization

Previous data demonstrate that L-type voltage-gated calcium channel (VGCC) blockers, which bind to α₁ subunits of VGCC to suppress Ca²⁺ entry into cells, inhibit the development of psychological dependence on drugs of abuse, suggesting the upregulation of L-type VGCC in the development of psychological dependence. However, there are few available data on changes of the auxiliary subunit α₂/δ modifying L-type VGCC under such conditions. We therefore investigated here the role of α₂/δ subunits of VGCCs in the brain of mouse after repeated treatment with morphine. The treatment with morphine increased α₂/δ subunit expression in the frontal cortex and the limbic forebrain of mice showing rewarding effect and sensitization to hyperlocomotion by morphine. The morphine-induced behavioral sensitization and place preference were also suppressed by gabapentin, which binds to an exofacial epitope of the α₂/δ auxiliary subunits of VGCCs. These findings indicate that the upregulation of α₂/δ subunit as well as α₁ subunits of VGCC in the frontal cortex and the limbic forebrain plays a critical role in development of morphine-induced rewarding effect and behavioral sensitization following neuronal plasticity.     

β1-Adrenoceptor distribution in the rat brain: An immunohistochemical study

Current knowledge of the central nervous system distribution of the β₁-adrenergic receptors (β₁-AR) is incomplete. Here we present a general map of the β₁-AR distribution in the rat brain. β₁-AR-immunoreactivity was detected throughout the entire rat brain, but particularly dense staining was observed in the cerebellar cortex and basal ganglia. Brainstem areas displaying significant β₁-AR-immunoreactivity include the ventrolateral medulla, nucleus ambiguus and the nucleus of the solitary tract. Within the hypothalamus, only the paraventricular nucleus and the median eminence (ME) showed β₁-AR immunostaining. Numerous β₁-AR-immunoreactive cells were also found in the hippocampus, basal ganglia and cerebral cortex. These results extend our knowledge of the expression profile of β₁-AR in the central nervous system. The identification of several distinct β₁-AR immunoreactive substrates linked with neuropathophysiological roles in cardiovascular disease supports the hypothesis that the therapeutic benefit of β₁-AR blockade may be conferred at least in part through central nervous system mechanisms.     

β subunits determine the time course of desensitization in rat α3 neuronal nicotinic acetylcholine receptors

Standard two-electrode voltage-clamp techniques were used to investigate some of the pharmacological and functional properties of two types of rat neuronal nicotinic acetylcholine receptors expressed in Xenopus oocytes after pairwise injection of 34 or 32 mRNAs. Currents of several A amplitude were elicited by fast application of micromolar concentrations of either acetylcholine (ACh) or 1,1-dimethyl-4-piperazine (DMPP). The activation of either receptor type by DMPP showed cooperativity (Hill coefficient, n1.7) with a half-maximal activation concentration (EC₅₀) of 15–30 M. In 34 receptors, ACh displayed cooperativity (n=1.8) but was less efficacious than DMPP, yet its EC₅₀ was about equal to that of DMPP. Finally, in 32 receptors, ACh was much less efficacious and had a much lower EC₅₀. Desensitization induced by either DMPP or ACh was slow in 34 nicotinic ACh receptors but was rapid and extensive in 32 receptors, causing a significant proportion of the response to wane within the first few seconds of agonist application.     

Distribution of metabotropic glutamate receptors in the parabrachial and Kölliker-Fuse nuclei of the rat

In the present study, we analysed the distribution and cellular localization of metabotropic glutamate receptors (1alpha, 2/3, 5) in parabrachial and K?lliker-Fuse nuclei using subtype-specific antisera. Immunolabelling revealed that different nuclei express different sets of metabotropic glutamate receptors. Metabotropic glutamate receptor la immunoreactivity was found in the K?lliker-Fuse nucleus and in several parabrachial nuclei, including the waist area, lateral crescent, medial, external medial and ventral lateral nuclei. The external lateral and internal lateral parabrachial nuclei were devoid of metabotropic glutamate receptor 1alpha immunoreactivity. Metabotropic glutamate receptor 5 immunoreactivity was observed in the K?lliker-Fuse and in the medial parabrachial nuclei, while in the remaining nuclei the staining was very weak. Again, the external lateral nucleus was devoid of metabotropic glutamate receptor 5 immunoreactivity. The metabotropic glutamate receptor 2/3 antisera stained all lateral parabrachial nuclei as well as the K?lliker-Fuse nucleus, while staining in the medial parabrachial nucleus was weak. Metabotropic glutamate receptor 1alpha immunoreactivity was observed on presumed dendritic profiles, while metabotropic glutamate receptor 5 immunoreactivity was found predominantly on neuronal cell bodies. Metabotropic glutamate receptor 2/3 immunoreactivity was present as a fine, punctate immunostaining in the neuropil. Our data suggest that glutamate release in the parabrachial and K?lliker-Fuse nuclei might induce a variety of second messenger cascades, as indicated by the presence or absence of certain types of metabotropic glutamate receptors.     

Gi2 proteins couple somatostatin receptors to low-conductance K+ channels in rat pancreatic α-cells

Somatostatin hyperpolarized rat pancreatic alpha-cells and inhibited spontaneous electrical activity by activating a low-conductance K+ channel (0.9 pS with physiological ionic gradients). This channel was insensitive to tolbutamide (a blocker of ATP-sensitive K+ channels) and apamin (an inhibitor of small-conductance Ca(2+)-activated K+ channels). Channel activation was prevented by pre-treating the cells with pertussis toxin, indicating the involvement of G-proteins. A direct interaction between an inhibitory G-protein and the somatostatin-activated K+ channel is suggested by the finding that intracellular application of guanosine 5'-O-(3-thiotriphosphate) (GTP gamma-S) and the G beta gamma subunit of G-proteins resulted in a transient stimulation of the current. Activation of the K+ current by somatostatin was inhibited by intracellular dialysis with specific antibodies to Gi1/2 and was not seen in cells treated with antisense oligonucleotides against G-proteins of the subtype Gi2. We conclude that somatostatin suppresses alpha-cell electrical activity by a Gi2-protein-dependent mechanism, which culminates in the activation of a sulphonylurea- and apamin-insensitive low-conductance K+ channel.     

Activation of peripheral κ/δ opioid receptors mediates 15-deoxy--prostaglandin J2 induced-antinociception in rat temporomandibular joint

This study assessed the effect of the agonist 15d-PGJ₂ administered into the rat temporomandibular joint (TMJ) on nociceptive behavioral and the anti-inflammatory potential of this prostaglandin on TMJ. It was observed that 15-deoxy-^Δ12,14-prostaglandin J₂ (15d-PGJ₂) significantly reduced formalin-induced nociceptive behavior in a dose dependent manner, however injection of 15d-PGJ₂ into the contralateral TMJ failed to reduce such effects. This antinociceptive effect is dependent on peroxisome proliferator-activated receptors-γ (PPAR-γ) since pre-treatment with GW9662 (PPAR-γ receptor antagonist) blocked the antinociceptive effect of 15d-PGJ₂ in the TMJ. In addition, the antinociceptive effect of 15d-PGJ₂ was also blocked by naloxone suggesting the involvement of peripheral opioids in the process. Confirming this hypothesis pre-treatment with κ, δ, but not μ receptor antagonists significantly reduced the antinociceptive effect of 15d-PGJ₂ in the TMJ. Similarly to opioid agonists, the 15d-PGJ₂ antinociceptive action depends on the nitric oxide (NO)/guanilate cyclase (cGMP)/ATP-sensitive potassium channel blocker(K⁺_ATP) channel pathway since it was prevented by the pre-treatment with the inhibitors of nitric oxide synthase (NOS; aminoguanidine), cGMP (ODQ), or the K⁺_ATP (glibenclamide). In addition, 15d-PGJ₂ (100 ng/TMJ) inhibits 5-HT-induced TMJ hypernociception. Besides, TMJ treated with 15d-PGJ₂ showed lower vascular permeability, assessed by Evan's Blue extravasation, and also lower neutrophil migration induced by carrageenan administration. Taken together, these results demonstrate that 15d-PGJ₂ has a potential peripheral antinociceptive and anti-inflammatory effect in the TMJ via PPAR-γ activation. The results also suggest that 15d-PGJ₂ induced-peripheral antinociceptive response in the TMJ is mediated by κ/δ opioid receptors by the activation of the intracellular l-arginine/NO/cGMP/K⁺_ATP channel pathway. The pharmacological properties of the peripheral administration of 15d-PGJ₂ highlight the potential use of this PPAR-γ agonist on TMJ inflammatory pain conditions.     

Stimulation of α1-adrenoceptors reduces glutamatergic synaptic input from primary afferents through GABAA receptors and T-type Ca channels

Activation of the descending noradrenergic system inhibits nociceptive transmission in the spinal cord. Although both α₁- and α₂-adrenoceptors in the spinal cord are involved in the modulation of nociceptive transmission, it is not clear how α₁-adrenoceptors regulate excitatory and inhibitory synaptic transmission at the spinal level. In this study, inhibitory and excitatory postsynaptic currents (IPSCs and EPSCs, respectively) were recorded from lamina II neurons in rat spinal cord slices. The specific α₁-adrenoceptor agonist phenylephrine significantly increased the frequency of GABAergic spontaneous IPSCs in a concentration dependent manner, and this effect was abolished by the α₁-adrenoceptor antagonist 2-(2,6-dimethoxyphenoxy)ethylaminomethyl-1,4-benzodioxane (WB4101). Phenylephrine also significantly reduced the amplitude of monosynaptic and polysynaptic EPSCs evoked from primary afferents. The inhibitory effect of phenylephrine on evoked monosynaptic glutamatergic EPSCs was largely blocked by the GABA_A receptor antagonist picrotoxin and, to a lesser extent, by the GABA_B receptor antagonist CGP55845. Furthermore, blocking T-type Ca²⁺ channels with amiloride or mibefradil diminished the inhibitory effect produced by phenylephrine or the GABA_A receptor agonist muscimol on monosynaptic EPSCs evoked from primary afferents. Collectively, these findings suggest that activation of α₁-adrenoceptors in the spinal cord increases synaptic GABA release, which attenuates glutamatergic input from primary afferents mainly through GABA_A receptors and T-type Ca²⁺ channels. This mechanism of presynaptic inhibition in the spinal cord may be involved in the regulation of nociception by the descending noradrenergic system.     

Association between α1-antitrypsin and bronchiectasis in patients with humoral immunodeficiency receiving gammaglobulin infusions

Background

In patients with humoral immunodeficiency, the progression of bronchiectasis has been known to occur despite adequate gammaglobulin therapy and in the absence of recurrent infections. This observation suggests that factors other than gammaglobulin replacement might play a part in the prevention of lung damage in this population. α₁-Antitrypsin deficiency can be associated with bronchiectasis, a chronic inflammatory lung disease. The protective levels of α₁-antitrypsin and phenotype in preventing bronchiectasis have not been thoroughly studied in the immunodeficient population. We hypothesized that patients with humoral immunodeficiencies on gammaglobulin infusions and bronchiectasis have lower median levels, but not necessary “classically” deficient levels, of α₁-antitrypsin compared with those without bronchiectasis.

Expression of sodium channel α- and β-subunits in the nervous system of themyelin-deficient rat

Summary Using subtype-specific riboprobes and a non-isotopein situ hybridization technique, the pattern of expression of the mRNAs for voltage dependent sodium channel -subunits I, II, III and NaG, and the ₁-subumt were compared inmyelin-deficient rats and unaffected male littermates. Tissues examined included the hippocampus, cerebellum, spinal cord and dorsal root ganglia. Previous studies have demonstrated that the expression of sodium channel - and ₁-subunits follows a distinct temporal and spatial pattern during development, characterized in part by greater expression of -subunit III and its mRNA during development than in the adult. We examined animals of 20–22 days of age, a time when, according to earlier reports, the unaffected animals should nearly have reached an adult expression pattern. Normal male littermates were indeed found to express a sodium channel subunit mRNA pattern generally consistent with previous reports on adult rats.Myelin-deficient animals exhibited an expression pattern identical to the unaffected littermates, indicating that myelination is not required for the progression from the embryonic to the adult expression pattern of sodium channel subunits.     

Integrin α2β1 targeted GdVO4:Eu ultrathin nanosheet for multimodal PET/MR imaging

Multifunctional nanoprobes open exciting possibilities for accurate diagnosis and therapy. In this research, we developed a ⁶⁴Cu-labeled GdVO₄:4%Eu two-dimension (2D) tetragonal ultrathin nanosheets (NSs) that simultaneously possess radioactivity, fluorescence, and paramagnetic properties for multimodal imaging. The carboxyl-functionalized Eu³⁺-doped GdVO₄ NSs were synthesized by a facile solvothermal reaction, followed by ligand exchange with polyacrylic acid (PAA). With ultrathin thickness of ∼5 nm and width of ∼150 nm, the carboxyl-functionalized NSs were further modified by DOTA chelator for ⁶⁴Cu labeling and Asp-Gly-Glu-Ala (DGEA) peptide for integrin α₂β₁ targeting. After initial evaluation of the cytotoxicity and targeting capability with PC-3 cells, the obtained multifunctional nanoprobes (⁶⁴Cu-DOTA-GdVO₄:4%Eu-DGEA) were further explored for targeted positron emission tomography (PET) and T₁-weighted magnetic resonance imaging (MRI) of PC-3 tumor (prostate cancer, high integrin α₂β₁ expression) in vivo. Based on the strong fluorescence of the NSs, the particle distribution in mouse tissues was also determined by fluorescent microscopy. In summary, GdVO₄:4%Eu NS is a potential multimodal multiscale nanoprobe that could not only be used for in vivo imaging, but also be tracked in cellular scale and ex vivo due to its fluorescent property.     

Effects of amyloid-β peptides on the serotoninergic 5-HT1A receptors in the rat hippocampus

A recent [¹⁸F]MPPF-positron emission tomography study has highlighted an overexpression of 5-HT_1A receptors in the hippocampus of patients with mild cognitive impairment compared to a decrease in those with Alzheimer's disease (AD) [Truchot, L., Costes, S.N., Zimmer, L., Laurent, B., Le Bars, D., Thomas-Antérion, C., Croisile, B., Mercier, B., Hermier, M., Vighetto, A., Krolak-Salmon, P., 2007. Up-regulation of hippocampal serotonin metabolism in mild cognitive impairment. Neurology 69 (10), 1012-1017]. We used in vivo and in vitro neuroimaging to evaluate the longitudinal effects of injecting amyloid-β (Aβ) peptides (1-40) into the dorsal hippocampus of rats. In vivo microPET imaging showed no significant change in [¹⁸F]MPPF binding in the dorsal hippocampus over time, perhaps due to spatial resolution. However, in vitro autoradiography with [¹⁸F]MPPF (which is antagonist) displayed a transient increase in 5-HT_1A receptor density 7 days after Aβ injection, whereas [¹⁸F]F15599 (a radiolabelled 5-HT_1A agonist) binding was unchanged suggesting that the overexpressed 5-HT_1A receptors were in a non-functional state. Complementary histology revealed a loss of glutamatergic neurons and an intense astroglial reaction at the injection site. Although a neurogenesis process cannot be excluded, we propose that Aβ injection leads to a transient astroglial overexpression of 5-HT_1A receptors in compensation for the local neuronal loss. Exploration of the functional consequences of these serotoninergic modifications during the neurodegenerative process may have an impact on therapeutics targeting 5-HT_1A receptors in AD.     

Ca-EGTA affects the relationship between [Ca2+] and tension in α-toxin permeabilized rat anococcygeus smooth muscle

Summary The relationship between calcium concentration ([Ca²⁺]) and force in smooth muscle can be studied by permeabilizing the sarcolemma and bathing the preparation in a mock intracellular solution. Normally [Ca²⁺] is set in these solutions using the Ca²⁺ chelator EGTA in the concentration range of 4–10 mm. This study shows that lowering total EGTA concentration ([EGTA]_t) below 10 mm depresses Ca²⁺-activated force generated in 0.1 m Ca²⁺. The observed threshold for the effect of EGTA_t is 0.2 mm, and the effect is maximal at approximately 10 mm. BAPTA, another Ca²⁺ chelator, also produces this effect. Tension production in smooth muscle is controlled by acto-myosin interaction. This in turn is mediated by the relative activities of myosin light chain kinase (MLCK) and phosphatase (MLCP). Inhibiting MLCP with Microcystin LR (10 m), an increase [EGTA_t] from 0.2 mm to 10 mm still enhanced force. This suggests that EGTA promotes phosphorylation of myosin by the activation of MLCK and not by inhibition of MLCP.