Natriorexigenic effect of baclofen is reduced by AT(1) receptor blockade in the lateral parabrachial nucleus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Esthetic Interim Acrylic Resin Prosthesis Reinforced with Metal Casting

Fabrication of an interim prosthesis is an important procedure in oral rehabilitation because it aids in determining the esthetics, phonetics, and occlusal relationship of the definitive restoration. The typical material (acrylic resin) used in interim prostheses commonly fails due to fractures. During extended oral rehabilitation with fixed partial prostheses, high strength interim prostheses are often required to protect hard and soft tissues, avoid dental mobility, and to allow the clinician and patient a chance to evaluate cosmetics and function before the placement of the definitive prosthesis. Furthermore, a satisfactory interim prosthesis can serve as a template for the construction of the definitive prosthesis. The maintenance of this prosthesis is important during treatment for protection of teeth and occlusal stability. Procedures to reinforce interim prostheses help to improve performance and esthetics in long-term treatment. Due to the low durability of acrylic resin in long-term use, the use of reinforcing materials, such as metal castings or spot-welded stainless steel matrix bands, is indicated in cases of extensive restoration and long-term treatment. This paper describes an easy technique for fabricating a fixed interim prosthesis using acrylic resin and a cast metallic reinforcement.

Análise radiográfica computadorizada da reabsorção radicular apical após a utilização de duas mecânicas ortodônticas

OBJETIVO: a reabsorção radicular apical é uma condição comumente observada durante e após o tratamento ortodôntico e fatores como tipo de aparelhagem ortodôntica utilizada, magnitude das forças aplicadas e duração do tratamento podem estar relacionados ao processo de arredondamento do ápice radicular. Buscou-se avaliar, por meio de imagens radiográficas computadorizadas, a quantidade de reabsorção no ápice radicular, quando da utilização de duas diferentes técnicas de mecânica ortodôntica fixa: Edgewise com acessórios padrão e Edgewise com acessórios totalmente programados. METODOLOGIA: a amostra constituiu-se de 20 pacientes tratados pelo mesmo profissional na Clínica de Ortodontia da Faculdade de Odontologia de Araçatuba - UNESP. Os pacientes foram divididos em dois grupos: grupo 1 (Tratados com Edgewise com acessórios padrão e fios de aço) e grupo 2 (Tratados com Edgewise com acessórios totalmente programados e fios de níquel-titânio). A avaliação radiográfica digital foi realizada por um único operador por meio de exposições radiográficas digitais feitas no início do tratamento ortodôntico, após retração de caninos e final de tratamento, utilizando o sistema de escores proposto por Levander e Malmgren (1988). RESULTADOS E CONCLUSÕES: a análise dos escores permitiu concluir que o tratamento ortodôntico empregando a técnica Edgewise com acessórios totalmente programados e fios de níquel-titânio apresentou menores graus de reabsorção radicular apical, em comparação à técnica Edgewise com acessórios padrão e fios de aço. Observou-se que, independentemente da técnica empregada, o tratamento ortodôntico como um todo apresentou um grau moderado de reabsorção radicular apical.

Orthodontic movement of teeth with intraosseous defects: histologic and histometric study in dogs

The purpose of this study was to evaluate histologically, in dogs, the periodontal healing of 1-walled intraosseous defects in teeth that were subjected to orthodontic movement toward the defects. The defects were surgically created bilaterally at the mesial aspects of the maxillary second premolars and distal aspects of the mandibular second premolars of 4 mongrel dogs. One week after creating the defects, an orthodontic appliance was installed, and the teeth were randomly assigned to 1 of 2 treatment groups: those in the test group received a titanium-molybdenum alloy rectangular wire spring that performed a controlled tipping root movement, and those in the control group received a passive stainless steel wire. Active orthodontic movement of the test teeth lasted 2 months and was followed by a stabilization period of another 2 months, after which the animals were killed. Throughout the study, routine daily plaque control was performed on the dogs with a topical application of a 2% chlorhexicline gel. The results showed no difference between the groups, with some regularization of the defects and periodontal regeneration limited to the apical portion of the defects. Histometric analysis showed a significant difference in bone height; on average, it was 0.53 mm smaller in the test group. It was concluded that orthodontic movement does not interfere with the healing of 1-walled intraosseous defects, with the exception of the linear extent of new bone apposition.

Inhibition of pilocarpine-induced salivation in rats by central noradrenaline

Peripheral treatment with cholinergic or adrenergic agonists results in salivation and the possibility of synergy between cholinergic and adrenergic efferent mechanisms in the control of salivation has been proposed. Central injections of the cholinergic agonist pilocarpine also induce salivation, while the effects of central injections of noradrenaline (norepinephrine) are not known. Here (a) the effects of intracerebroventricular (icv) injection of noradrenaline on the salivation induced by icv or intraperitoneal (i.p.) injection of pilocarpine and (b) the receptors involved in the effects of central noradrenaline on pilocarpine-induced salivation were investigated. Male Holtzman rats with a stainless-steel guide cannula implanted into the lateral ventricle were used. Rats were anaesthetized with tribromoethanol (200 mg/kg body weight) and saliva was collected on small, preweighed cotton balls inserted into the animal's mouth. Noradrenaline (40, 80 and 160 nmol/l mul) injected icv reduced the salivary secretion induced by pilocarpine (0.5 mumol/l mul) injected icv. Noradrenaline (80 and 160 nmol/l mul) injected icv also reduced the salivation induced by pilocarpine (4 mumol/kg) injected i.p. Previous treatment with the alpha(2)-adrenergic receptor antagonists RX 821002 (40, 80 and 160 nmol/l mul) or yohimbine (160 and 320 nmol/l mul) abolished the inhibitory effect produced by icv injection of noradrenaline on pilocarpine-induced salivation in rats. Prazosin (alpha(1)-adrenergic receptor antagonist) injected icv did not change the effect of noradrenaline on pilocarpine-induced salivation. Prior icv injection of only RX 821002 (80 or 160 nmol/l mul) or yohimbine (320 nmol/l mul) increased pilocarpine-induced salivation. The results show that (1) contrary to its peripheral effects, noradrenaline acting centrally inhibits cholinergic-induced salivation in rats; (2) central mechanisms involving alpha(2)-adrenergic receptors inhibit pilocarpine-induced salivation. (C) 2002 Elsevier B.V. Ltd. All rights reserved.

Cardiovascular responses to microinjection of L-glutamate into the NTS in AV3V-lesioned rats

The excitatory amino acid L-glutamate injected into the nucleus of the solitary tract (NTS) in unanesthetized rats similar to peripheral chemoreceptor activation increases mean arterial pressure (MAP) and reduces heart rate. In this study, we investigated the effects of acute (I day) and chronic (15 days) electrolytic lesions of the preoptic-periventricular tissue surrounding the anteroventral third ventricle (AV3V region) on the pressor and bradycardic responses induced by injections of L-glutamate into the NTS or peripheral chemoreceptor activation in unanesthetized rats. Male Holtzman rats with sham or electrolytic AV3V lesions and a stainless steel cannula implanted into the NTS were used. Differently from the pressor responses (28 +/- 3 mm Hg) produced by injections into the NTS of sham-lesioned rats, L-glutamate (5 nmol/ 100 nl) injected into the NTS reduced MAP (-26 +/- 8 mm Hg) or produced no effect (2 7 turn Hg) in acute and chronic AV3V-lesioned rats, respectively. The bradycardia to L-glutamate into the NTS and the cardiovascular responses to chemoreflex activation with intravenous potassium cyanide or to baroreflex activation with intravenous phenylephrine or sodium nitroprusside were not modified by AV3V lesions. The results show that the integrity of the AV3V region is essential for the pressor responses to L-glutamate into the NTS but not for the pressor responses to chemoreflex activation, suggesting dissociation between the central mechanisms involved in these responses. (C) 2004 Elsevier B.V. All rights reserved.

Vasopressin and angiotensin receptors of the medial septal area of the brain in the control of thirst and salt appetite induced by vasopressin in water-deprived and sodium-depleted rats

In this study we investigated the influence of d(CH2)(5)-Tyr (Me)-AVP (A(1) AVP) and [Adamanteanacatyl(1),D-ET-D-Tyr(2), Va1(4), aminobutyril(6) ,As-8,As-9]-AVP 9 (A(2)AVP), antagonists of V-1 and V-2 arginine(8)-vasopressin (AVP) receptors, respectively, as well as the effects of losartan and CGP42112A, antagonists of angiotensin II (ANGII) AT(1) and AT(2), receptors, respectively, on water and 0.3 M sodium intake induced by water deprivation or sodium depletion (furosemide treatment) and enhanced by AVP injected into the medial septal area (N4SA). A stainless steel carmulawas implanted into the medial septal area (NISA) of male Holtzman rats AVP injection enhanced water and sodium intake in a dose-dependent manner. Pretreatment with V-1 antagonist injected into the MSA produced a dose-dependent reduction, whereas prior injection of V-2 antagonist increased, in a dose-dependent manner, the water and sodium responses elicited by the administration of AVP. Both AT(1) and AT(2) antagonists administered into the MSA elicited a concentration-dependent decrease in water and sodium intake induced by AVP, while simultaneous injection of the two antagonists was more effective in decreasing AVP responses. These results also indicate that the increase in water and sodium intake induced by AvT was mediated primarily by MSA AT(1) receptors. (c) 2007 Published by Elsevier B.V.

Central alpha(2) adrenergic receptors and cholinergic-induced salivation in rats

Salivation induced by intraperitoneal (i.p.) injections of pilocarpine (cholinergic agonist) is reduced by intracerebroventricular (i.c.v.) injections of moxonidine (alpha(2) adrenergic and imidazoline receptor agonist). In the present study, we investigated the involvement of central alpha(2) adrenergic receptors in the inhibitory effect of i.c.v. moxonidine on i.p. pilocarpine-induced salivation. Male Holtzman rats with stainless steel cannula implanted into the lateral ventricle (LV) were used. Saliva was collected using pre-weighted small cotton balls inserted into the animal's mouth under ketamine (100 mg kg(-1)) anesthesia. Salivation was induced by i.p. injection of pilocarpine (4 mu mol kg(-1)). Pilocarpine-induced salivation was reduced by i.c.v. injection of moxonidine (10 nmol) and enhanced by i.c.v. injections of either RX 821002 (160 nmol) or yohimbine (320 nmol). The inhibitory effect of i.c.v. moxonidine on pilocarpine-induced salivation was abolished by prior i.c.v. injections of the alpha(2) adrenergic receptor antagonists, RX 821002 (160 nmol) or yohimbine (160 and 320 nmol). The alpha(1) adrenergic receptor antagonist prazosin (320 nmol) injected i.c.v. did not change the effect of moxonidine on pilocarpine-induced salivation. The results suggest that moxonidine acts on central alpha(2) adrenergic receptors to inhibit pilocarpine-induced salivation, and that this salivation is tonically inhibited by central alpha(2) adrenergic receptors. (C) 2002 Elsevier B.V. All rights reserved.

Central blockade of nitric oxide synthesis reduces moxonidine-induced hypotension

1 Nitric oxide (NO) and alpha(2)-adrenoceptor and imidazoline agonists such as moxonidine may act centrally to inhibit sympathetic activity and decrease arterial pressure.2 In the present study, we investigated the effects of pretreatment with L-NAME ( NO synthesis inhibitor), injected into the 4th ventricle (4th V) or intravenously (i.v.), on the hypotension, bradycardia and vasodilatation induced by moxonidine injected into the 4th V in normotensive rats.3 Male Wistar rats with a stainless steel cannula implanted into the 4th V and anaesthetized with urethane were used. Blood flows were recorded by use of miniature pulsed Doppler flow probes implanted around the renal, superior mesenteric and low abdominal aorta.4 Moxonidine (20 nmol), injected into the 4th V, reduced the mean arterial pressure (-42+/-3 mmHg), heart rate (-22+/-7 bpm) and renal (-62+/-15%), mesenteric (-41+/-8%) and hindquarter (-50+/-8%) vascular resistances.5 Pretreatment with L-NAME (10 nmol into the 4th V) almost abolished central moxonidine-induced hypotension (-10+/-3 mmHg) and renal (-10+/-4%), mesenteric (-11+/-4%) and hindquarter (-13+/-6%) vascular resistance reduction, but did not affect the bradycardia (-18+/-8 bpm).6 the results indicate that central NO mechanisms are involved in the vasodilatation and hypotension, but not in the bradycardia, induced by central moxonidine in normotensive rats. British Journal of Pharmacology (2004).

Serotonergic mechanisms of the lateral parabrachial nucleus on DOCA-induced sodium intake

It has been shown that the serotonergic mechanisms of the lateral parabrachial nucleus (LPBN) inhibit NaCl intake in different models of angiotensin II (ANG II)-dependent NaCl intake in rats. However, there is no information about the involvement of LPBN serotonergic mechanisms on NaCl intake in a model of NaCl intake not dependent on ANG II like deoxycorticosterone (DOCA)-induced NaCl intake. Therefore, in this study we investigated the effects of bilateral injections of serotonergic agonist and antagonist into the LPBN on DOCA-induced 1.8% NaCl intake in rats. Male Holtzman rats were treated with s.c. DOCA (10 mg/rat each every 3 days). After a period of training, in which the rats had access to 1.8% NaCI during 2 h for several days, the rats were implanted with stainless steel cannulas bilaterally into the LPBN. Bilateral injections of the serotonergic receptor antagonist methysergide (4 mug/0.2 mul each site) in the LPBN increased 1.8% NaCI intake (32.2+/-3.9 versus vehicle: 15.0+/-1.6 ml/2 h, n = 10) and water intake (11.5+/-3.5 versus vehicle: 3.2+/-1.0 ml/2 h). Injections of the serotonergic 5HT(2A/2C) receptor agonist DOI (5 mug/0,2 mul each site) in the LPBN reduced 1.8% NaCl intake (6.8+/-1.7 versus saline: 12.4+/-1.9 ml/2 h, n = 10) and water intake (2.2+/-0.8 versus saline: 4.4+/-1.0 ml/2 h). Besides the previously demonstrated importance for the control of ANG II-dependent water and NaCl intake, the data show that the serotonergic inhibitory mechanisms of the LPBN are also involved in the control of DOCA-induced NaCl intake. (C) 2000 Elsevier B.V. B.V. All rights reserved.

Role of nitric oxide and beta-adrenoceptors of the central nervous system on the salivary flow induced by pilocarpine injection into the lateral ventricle

Our studies have focused on the effect of L-NG-nitroarginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase (NOS), and L-arginine, the substrate of NOS, on salivary secretion induced by the administration of pilocarpine into the lateral cerebral ventricle (LV) of rats. The present study has also investigated the role of the beta-adrenergic agonists and antagonist injected into LV on the salivary secretion elicited by the injection of pilocarpine into LV. Male Holtzmann rats with a stainless-steel cannula implanted into the LV were used. The amount of salivary secretion was studied over a 7-min period after injection of pilocarpine, isoproterenol, propranolol, salbutamol, salmeterol, L-NAME and L-arginine. The injection of pilocarpine (10, 20, 40, 80 and 160 mug/mul) into LV produced a dose-dependent increase in salivary secretion. The injection of L-NAME (40 mug/mul) into LV alone produced an increase in salivary secretion. The injection of L-NAME into LV previous to the injection of pilocarpine produced an increase in salivary secretion. L-Arginine (30 mug/mul) injected alone into LV produced no change in salivary secretion. L-Arginine injected into LV attenuated pilocarpine-induced salivary secretion. The isoproterenol (40 nmol/mul) injected into LV increased into LV increased the salivary secretion. When injected previous to pilocarpine at a dose of 20 and 40 mug/mul, isoproterenol produced and additive effect on pilocarpine-induced salivary secretion. The 40-nmol/mul dose of propranolol injected alone or previous to pilocarpine into LV attenuated the pilocarpine-induced salivary secretion. The injection of salbutamol (40 nmol/mul), a specific beta-2 agonist, injected alone into LV produced no change in salivary secretion and when injected previous to pilocarpine produced and increase in salivary secretion. The 40-nmol/mul dose of salmeterol, a long-acting beta-2 agonist, injected into LV alone or previous to pilocarpine produced no change in salivary secretion. The results have shown that central injections of L-NAME and L-arginine interfere with the salivary secretion, which implies that might participate in pilocarpine-induced salivary secretion. The interaction between cholinergic and beta-adrenergic receptors of the central nervous system (CNS) for the control of salivary secretion can also be postulated. (C) 2002 Elsevier B.V. All rights reserved.

Moxonidine and rilmenidine injected into the medial septal area reduces the salivation induced by pilocarpine

We determined the effects of moxonidine and rilmenidine 20 mol (alpha(2)-adrenergic and imidazoline receptor agonists) injected into the medial septal area (MSA) on the pilocarpine-induced salivation, when injected intraperitoneally (i.p.), of male Holtzman rats weighing 250300 g, with stainless-steel cannula implanted into the MSA. The rats were anesthetized with zoletil 50 mg kg(-1) b.wt. (tiletamine chloridrate 125.0 mg and zolazepan chloridrate 125.0 mg) into quadriceps muscle intramuscularly (IM), saliva was collected using pre-weighed small cotton balls inserted in the animal's mouth. The pre-treatment with moxonidine injected into the MSA reduced the salivation induced by pilocarpine (1 mg kg(-1)) injected i.p. (12 +/- 3 mg min(-1)) vs. control (99 +/- 9 mg min(-1)). The pre-treatment with rilmenidine 40 nmol also reduced the salivation induce by pilocarpine injected i.p. (20 +/- 5 mg min(-1)) vs. control (94 +/- 7 mg min(-1)). Idazoxan 40 nmol (imidazoline receptor antagonist) injected into the MSA previous to moxonidine and rilmenidine partially blocked the effect of moxonidine and totally blocked the rilmenidine effect in pilocarpine-induced salivation injected i.p. (60 +/- 8 and 95 +/- 10 mg min(-1), respectively). Yohimbine 40 nmol (alpha(2)-adrenergic receptor antagonist) injected into the MSA previously to moxonidine and rilmenidine partially blocked the moxonidine effect but produced no change on the rilmenidine effect on i.p. pilocarpine-induced salivation (70 +/- 6 and 24 +/- 6 mg min(-1), respectively). Injection of these alpha(2)-adrenergic and imidazoline agonists and antagonists agents i.p. produced no change on i.p. pilocarpine-induced salivation. These results show that central, but not peripheral, injection of alpha(2)-adrenergic and imidazoline agonists' agents inhibit pilocarpine-induced salivation. Idazoxan, an imidazoline receptor antagonist, totally inhibits the rilmenidine effect and partially inhibits the moxonidine effect on pilocarpine-induced salivation. Yohimbine produced no change on rilmenidine effect but partially inhibited the moxonidine effect. Both of these antagonists when injected into the MSA previous to pilocarpine i.p. potentiated the sialogogue effect of pilocarpine. The results suggest that alpha(2)-adrenergic/imidazoline receptor of the MSA when stimulated blocked pilocarpine-induced salivation in rats when injected intraperitonially These receptors of the medial septal area have an inhibitory mechanism on salivary secretion. (C) 2004 Elsevier B.V. All rights reserved.

Alpha(2)-adrenergic activation in the lateral parabrachial nucleus induces NaCi intake under conditions of systemic hyperosmolarity

The inhibition of sodium intake by increased plasma osmolarity may depend on inhibitory mechanisms present in the lateral parabrachial nucleus. Activation of alpha(2)-adrenergic receptors in the lateral parabrachial nucleus is suggested to deactivate inhibitory mechanisms present in this area increasing fluid depletion-induced 0.3 M NaCl intake. Considering the possibility that lateral parabrachial nucleus inhibitory mechanisms are activated and restrain sodium intake in animals with increased plasma osmolarity, in the present study we investigated the effects on water and 0.3 M NaCl intake produced by the activation of alpha(2)-adrenergic receptors in the lateral parabrachial nucleus in rats with increased plasma osmolarity. Male Holtzman rats with stainless steel cannulas implanted bilaterally into the lateral parabrachial nucleus were used. One hour after intragastric 2 M NaCl load (2 ml), bilateral injections of moxonidine (alpha(2)-adrenergic/imidazoline receptor agonist, 0.5 nmol/0.2 mu l, n=10) into the lateral parabrachial nucleus induced a strong ingestion of 0.3 M NaCl intake (19.1 +/- 5.5 ml/2 h vs. vehicle: 1.8 +/- 0.6 ml/2 h), without changing water intake (15.8 +/- 3.0 ml/2 h vs. vehicle: 9.3 +/- 2.0 ml/2 h). However, moxonidine into the lateral parabrachial nucleus in satiated rats not treated with 2 M NaCl produced no change on 0.3 M NaCl intake. The pre-treatment with RX 821002 (alpha(2)-adrenergic receptor antagonist, 20 nmol/0.2 mu l) into the lateral parabrachial nucleus almost abolished the effects of moxonidine on 0.3 M NaCl intake (4.7 +/- 3.4 ml/2 h). The present results suggest that alpha(2)-adrenergic receptor activation in the lateral parabrachial nucleus blocks inhibitory mechanisms, thereby allowing ingestion of hypertonic NaCl under conditions of extracellular hyperosmolarity. We suggest that during cell dehydration, circuits subserving sodium appetite are activated, but at the same time strongly inhibited through the lateral parabrachial nucleus. (c) 2006 IBRO. Published by Elsevier Ltd. All rights reserved.

Functional relationship between subfrnical organ cholinergic stimulation and nitrergic activation influencing cardiovascular and body fluid homeostasis

We have studied the effects of L-NG-nitro arginine methyl esther (L-NAME), L-arginine (LAR), inhibitor and a donating nitric oxide agent on the alterations of salivary flow, water intake, arterial blood pressure (MAP) and heart rate (HR) induced by the injection pilocarpine into the subfornical organ (SFO). Rats (Holtzman 250-300 g) were anesthetized with 2, 2, 2-tribromoethanol (20 mg/100 kg b. wt.) and a stainless steel carmula were implanted into their SFO. The volume of injection was 0.2 mu l. The amount of saliva secretion was studied over a 5-min period. Pilocarpine (40 mu g), L-NAME (40 mu g) and LAR (30 mu g) were used in all experiments for the injection into the SFO. Pilocarpine (10, 20, 40, 80 and 160 mu g) injected into SFO elicited a concentration-dependent increase in salivary secretion. L-NAME injected prior to pilocarpine into the SFO increased salivary secretion and water intake due to the effect of pilocarpine. LAR injected prior to pilocarpine into the SFO attenuated the salivary secretion and water intake. Pilocarpine, injected into the SFO increased the MAP and decreased heart rate (HR). L-NAME injected prior to pilocarpine into the SFO potentiated the pressor effect of pilocarpine with a decrease in HR. LAR injected into the SFO prior to pilocarpine attenuated the increase in MAP with no changes in HR. The present study suggests that the SFO nitrergic cells interfere in the cholinergic pathways implicated in the control of salivary secretion, fluid and cardiovascular homeostasis. (c) 2007 Elsevier B.V All rights reserved.

Moxonidine reduces pilocarpine-induced salivation in rats

Cholinergic, agonists activate salivation and the alpha (2)-adrenergic and imidazoline receptor agonists induce opposite effects. In the present study, we investigated the effects of intracerebroventricular (i.c.v.) or intraperitoneal (i.p.) injection of moxonidine (an a-adrenergic and imidazoline receptor agonist) on the salivation induced by the cholinergic agonist pilocarpine. Male Holtzman rats wish stainless steel cannula implanted into the lateral ventricle (LV) were used. In rats anesthetized with tribromoethanol (200 mg kg(-1)), saliva was collected using pre-weighed small cotton balls inserted in the animal's mouth. The treatment with moxonidine (5, 10 and 20 nmol in 1 mul) injected,i.c.v. reduced the salivation induced by pilocarpine (1 mg kg(-1)) injected i.p. (48 +/- 5, 17 +/- 2 and 15 +/- 2 mg min(-1) vs. control, 73 +/- 7 mg min(-1)). The same doses of moxonidine injected i.c.v. also reduced the salivary secretion induced by pilocarpine (500 nmol in 1 mul). injected i.c.v. (44 +/- 1, 14 +/- 2 and 20 +/- 3 mg min(-1) vs. control, 51 +/- 2 mg min(-1)). Injection of moxonidine (20 nmol in 0.1 ml) i.p. produced no chance on i.p. pilocarpine-induced salivation (58 +/- 4 mg min(-1) vs. control, 50 +/- 4 mg min(-1)). The results show that central, but not peripheral, injection of moxonidine inhibit,. pilocarpine-induced salivation, suggesting that central mechanisms activated by alpha (2)-adrenergic/imidazoline agonists inhibit cholinergic-induced salivation in rats. (C) 2001 Elsevier B.V. B.V. All rights reserved.