RESSALVA Atendendo solicitação do(a) autor(a), o texto completo desta tese será disponibilizado somente a partir de 02/02/2025. UNIVERSIDADE ESTADUAL PAULISTA - UNESP CÂMPUS DE JABOTICABAL ADMINISTRAÇÃO INTRANASAL versus INTRAMUSCULAR DE AZAPERONE, MIDAZOLAM E CETAMINA EM SUÍNOS Isabela Peixoto Rabelo Médica Veterinária 2024 UNIVERSIDADE ESTADUAL PAULISTA - UNESP CÂMPUS DE JABOTICABAL ADMINISTRAÇÃO INTRANASAL versus INTRAMUSCULAR DE AZAPERONE, MIDAZOLAM E CETAMINA EM SUÍNOS Discente: Isabela Peixoto Rabelo Orientador: Prof. Dr. Carlos Augusto Araújo Valadão Tese apresentada à Faculdade de Ciências Agrárias e Veterinárias – Unesp, Câmpus de Jaboticabal, como parte das exigências para a obtenção do título de Doutor em Cirurgia Veterinária 2024 Sistema de geração automática de fichas catalográficas da Unesp. Biblioteca da Faculdade de Ciências Agrárias e Veterinárias, Jaboticabal. Dados fornecidos pelo autor(a). Essa ficha não pode ser modificada. R114a Rabelo, Isabela Peixoto Administração intranasal versus intramuscular de azaperone, midazolam e cetamina em suínos / Isabela Peixoto Rabelo. -- Jaboticabal, 2024 55 p. : il., tabs. Tese (doutorado) - Universidade Estadual Paulista (Unesp), Faculdade de Ciências Agrárias e Veterinárias, Jaboticabal Orientador: Carlos Augusto Araújo Valadão 1. Anestesia veterinária. 2. Suínos como animal de laboratório. 3. Medicação intranasal. I. Título. DADOS CURRICULARES DA AUTORA ISABELA PEIXOTO RABELO – nascida na cidade de São João del Rei, Minas Gerais, no dia 01 de junho de 1992. Médica Veterinária graduada pela Universidade Federal de Lavras (UFLA) em Lavras, MG, com início em março de 2010 e término em julho de 2015. Realizou estágio curricular no ano de 2015 na área de Cirurgia Veterinária de Grandes Animais no Hospital Veterinário da Faculdade de Medicina Veterinária e Zootecnia da USP em São Paulo, SP, e na Seção Veterinária do 1º Regimento da Cavalaria de Guardas do Exército Brasileiro em Brasília, DF. Concluiu Programa de Residência em Área Profissional da Saúde – Medicina Veterinária e Saúde, na subárea de Clínica Cirúrgica e Anestesiologia de Grandes Animais no Hospital Veterinário de Grandes Animais da UFLA, com início em março de 2016 e término em fevereiro de 2018, sob orientação da Professora Doutora Rosa Maria Cabral. Iniciou curso de mestrado em 2018 e, em 2019, obteve título de Mestre em Cirurgia Veterinária pela Universidade Estadual Paulista “Júlio de Mesquita Filho” UNESP - Câmpus de Jaboticabal, sob orientação do Professor Doutor Carlos Augusto Araújo Valadão. Atualmente é aluna de doutorado no Programa de Pós-graduação em Cirurgia Veterinária, com início em março de 2020, sob orientação do Professor Doutor Carlos Augusto Araújo Valadão. AGRADECIMENTOS Agradeço: Aos meus pais, Giovanna Cherfên Peixoto e José Maria Rabelo, por sempre acreditarem e confiarem nas minhas escolhas. Vocês são meu maior exemplo de vida. Amo muito vocês. À minha irmã e melhor amiga, Sara Peixoto Rabelo. Obrigada pela cumplicidade diária e por toda ajuda na construção desde trabalho. Te amo de todo coração. À minha madrinha, Lane Maria Rabelo, por todo amor e por ser um exemplo incrível de mulher na ciência. Te amo Ao meu orientador Prof. Dr. Carlos Augusto Araújo Valadão. Por ser, mais que um orientador, mas também um amigo. Obrigada pelo apoio, parceria e paciência ao longo de todos esses anos À minha irmãzinha que escolhi para vida, Ariane. Obrigada por não deixar a distância nos afastar e por estar sempre aqui por mim. Te amo. Aos meus pequenos: Lilith, Hel e Caramelo, por colorirem meus dias. Aos amigos e parceiros de experimento: Cinthya, Inácio, Vanessa, Ariadne. Este trabalho não teria acontecido se não fosse por vocês. Muito obrigada! Aos professores, técnicos e colegas da UNESP Jaboticabal, pela parceria direta e indireta neste experimento. Ao professor Luciano Hauschild e aos seus alunos, integrantes do LabSui, que generosamente nos abriram as portas do laboratório para realização deste trabalho. Obrigada por todo suporte e parceria. À FAPESP (RT processo nº 2020/09633-0) e CAPES (código 33004102.069- P8), pelo auxílio e pela bolsa de estudo, respectivamente, para que esse projeto fosse realizado com sucesso. À DESVET, pela parceria e fornecimento do Destress. Aos porquinhos que integraram este projeto. Minha eterna e mais sincera gratidão. À turminha da Sweet 14: Ana, Dani, Cin, Fogs, Pão e Pauli. Obrigada pela amizade, carinho e amor. Jaboticabal não seria a mesma sem vocês. À minha segunda família do Bernarda: Glau, Rafa, Bi, Mani, Ise, Amandinha, Ju, Dacota e Ti. Por toda confiança e acolhimento. Amo e admiro cada um. Estar com vocês é sempre um respiro, a melhor parte da minha semana. Aos meus demais amigos que estão longe, ou que não vejo tanto quanto eu gostaria. Felizmente, a lista é extensa. Mas saibam que penso com muito carinho em todos vocês. Obrigada por contribuírem com a construção da pessoa que me tornei. O presente trabalho foi realizado com apoio da Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES) - Código de Financiamento 001. i SUMÁRIO CERTIFICADO - CEUA ......................................................................................................... ii RESUMO ............................................................................................................................... iii ABSTRACT .......................................................................................................................... iv LISTA DE ABREVIATURAS .................................................................................................. v 1. INTRODUÇÃO ............................................................................................................... 7 2. REVISÃO DE LITERATURA .......................................................................................... 8 2.1. Suínos como modelo experimental da espécie humana ............................................. 8 2.2. Considerações sobre sedação e anestesia de suínos .............................................. 11 2.2.1. Características comportamentais, fisiológicas e anatômicas ................................ 11 2.2.2. Contenção física e manejo ................................................................................... 14 2.2.3. Vias de administração .......................................................................................... 15 2.2.4. Risco associado e monitoração anestésica .......................................................... 18 2.3. Sedação e contenção farmacológica de suínos ....................................................... 19 2.3.1. Azaperone ............................................................................................................ 19 2.3.2. Midazolam ............................................................................................................ 21 2.3.3. Cetamina .............................................................................................................. 23 3. MATERIAL E MÉTODOS ............................................................................................. 27 3.1. Animais e instalações ............................................................................................... 27 3.2. Delineamento experimental e formação dos grupos ................................................. 27 3.3. Administração intranasal e intramuscular ................................................................. 28 3.4. Coleta de sangue arterial (fase I) ............................................................................. 29 3.5. Parâmetros fisiológicos e comportamentais (fase II) ................................................ 29 3.6. Análise estatística .................................................................................................... 30 4. RESULTADOS ............................................................................................................. 31 4.1. Características da contenção química ...................................................................... 31 4.2. Parâmetros fisiológicos e gasométricos ................................................................... 32 5. DISCUSSÃO ................................................................................................................ 38 5.1. Características da contenção química ...................................................................... 38 5.2. Parâmetros fisiológicos e gasométricos ................................................................... 40 6. CONCLUSÃO .............................................................................................................. 42 7. REFERÊNCIAS ........................................................................................................... 42 ii CERTIFICADO - CEUA iii ADMINISTRAÇÃO INTRANASAL versus INTRAMUSCULAR DE AZAPERONE, MIDAZOLAM E CETAMINA EM SUÍNOS RESUMO – O objetivo deste estudo foi propor uma alternativa para a contenção química (CQ) de suínos, visando manter a homeostase ao minimizar o estresse da contenção. Para isso, foram comparadas as administrações intranasal e intramuscular da associação de azaperone (3 mg/kg), midazolam (0,3 mg/kg) e cetamina (7 mg/kg) (AMC) em 16 suínos adultos, machos imunocastrados, hígidos, de linhagem mista (Landrace x Large White), pesando 102 ± 12 kg. Na fase I, com duração de quatro dias, esses animais foram distribuídos aleatoriamente nos grupos intranasal (GIN, n = 8) e intramuscular (GIM, n = 8), para análise de gases e eletrólitos no sangue arterial aos 10, 20, 30, 45, 60, e 90 minutos após administração do AMC. A fase II foi realizada seis dias após o início da fase I. Os 16 suínos foram alocados em ambos os grupos (GIM, n = 16 / GIN, n = 16) e submetidos ao mesmo protocolo de CQ referido na fase I, com intervalo de 96 horas entre administrações. Nesta segunda fase, foram avaliados os parâmetros comportamentais (intensidade da CQ, relaxamento muscular, perda de reflexo postural e resposta ao estímulo sonoro) e vitais (frequência cardíaca – FC; frequência respiratória – fR; saturação periférica de oxigênio – SpO2 e temperatura retal – Tretal) logo após decúbito (Tdec) e aos 5, 15, 30, 45, 60 e 90 minutos após as administrações. Além disso, foi determinado o período de latência e duração da CQ. A latência para o decúbito foi menor no GIN (GIN: 63 ± 47 segundos; GIM: 113 ± 39 segundos; p = 0,002), assim como a duração da CQ (GIN: 119 ± 54 minutos; GIM: 163 ± 47; p = 0,0015). Os animais do GIN apresentaram maior grau de relaxamento muscular (p < 0,05). Observou-se uma taquicardia inicial, seguida por redução da FC de T5 a T90, nos dois tratamentos (p < 0,05). A fR elevou-se entre T45 e T90 no GIN, em relação ao Tdec. Houve redução da Tretal no GIM a partir de T45. Não foram detectadas alterações metabólicas severas, apenas elevação da PaCO2 no T90 no GIM (p < 0,05) e incidência de hipoxemia leve (entre 73 e 81 mmHg) em 47% dos animais do GIM. No GIN, a [Ca2+] reduziu com relação ao basal no T45. Em suma, a administração intranasal promoveu contenção química eficaz, com menor duração e período de latência, e mínimas alterações metabólicas e respiratórias. Palavras-chave: Benzodiazepínicos, tranquilização, modelo animal, sedação, tranquilizantes iv INTRANASAL vs INTRAMUSCULAR ADMINISTRATION OF AZAPERONE, MIDAZOLAM AND KETAMINE IN PIGS ABSTRACT – The aim of this study was to propose an alternative for the chemical restraint (CR) of swine, aiming to maintain homeostasis by minimizing the stress of restraint. We compared the intranasal and intramuscular administrations of the combination of azaperone (3 mg/kg), midazolam (0.3 mg/kg), and ketamine (7 mg/kg) (AMK) in 16 adult, immunocastrated male swine, healthy, of mixed lineage (Landrace x Large White), weighing 102 ± 12 kg. In Phase I, lasting four days, these animals were randomly allocated to the intranasal (GIN, n = 8) and intramuscular (GIM, n = 8) groups, for analysis of arterial blood gases and electrolytes at 10, 20, 30, 45, 60, and 90 minutes after AMK administration. Phase II was conducted six days after the start of Phase I. The 16 pigs were allocated to both groups (GIN, n = 16 / GIM, n = 16) and submitted to the same CR protocol referred in Phase I, with a 96-hour interval between administrations. In this second phase, behavioral (CR intensity, muscle relaxation, loss of postural reflex, and response to sound stimulus) and vital parameters (heart rate - HR; respiratory rate - RR; peripheral oxygen saturation - SpO2; and rectal temperature - Trect) were evaluated immediately after recumbency (Trec) and at 5, 15, 30, 45, 60, and 90 minutes after administrations. Additionally, the latency period and duration of CR were determined. The latency to recumbency was shorter in GIN (GIN: 63 ± 47 seconds; GIM: 113 ± 39 seconds; p = 0.002), as well as the duration of CR (GIN: 119 ± 54 minutes; GIM: 163 ± 47; p = 0.0015). Animals in the GIN showed a higher degree of muscle relaxation (p < 0.05). Initial tachycardia was observed, followed by a reduction in HR from T5 to T90 in both treatments (p < 0.05). RR increased between T45 and T90 in the GIN, compared to Trec. There was a reduction in Trect in GIM from T45 onwards. No severe metabolic changes were detected, only an increase in PaCO2 at T90 in GIM (p < 0.05) and mild hypoxemia (between 73 and 81 mmHg) in 47% of the animals in the GIM. In the GIN, [Ca2+] decreased compared to baseline at T45. In summary, intranasal administration promoted effective chemical restraint, with shorter duration and latency period, and minimal metabolic and respiratory changes. Keywords: Tranquilizers, benzodiazepines, sedation, animal model v LISTA DE ABREVIATURAS AMC: Azaperone (3 mg/kg) + Midazolam (0,3 mg/kg) + Cetamina (7 mg/kg) AMPA: α-amino-3-hidroxi-5-metil-4- isoxazolpropiônico BHE: Barreira hematoencefálica bpm: Batimentos por minuto Ca2+: Íon cálcio CEUA: Comissão de ética no uso de animais COVID-19: “Corona vírus disease” CQ: contenção química DC: Débito cardíaco EB: Excesso de bases fR: Frequência respiratória FC: Frequência cardíaca FCAV: Faculdade de Ciências Agrárias e Veterinárias GABA: Ácido γ-aminobutírico GIM: Grupo intramuscular GIN: grupo intranasal HCO3 -: Íon bicarbonato HM: Hipertermia maligna IM: Intramuscular IN: Intranasal IV: Intravenosa K+: íon potássio mpm: movimentos por minuto Na+: Íon sódio vi NMDA: N-Metil-D-Aspartato O2: oxigênio PaCO2: pressão arterial de dióxido de carbono PaO2: pressão arterial de oxigênio PAM: Pressão arterial média pH: potencial hidrogeniônico SARS-CoV-2: Coronavírus associado à síndrome respiratória aguda SNC: Sistema nervoso central T: tempo Tdec: Tempo do decúbito Tretal: Temperatura retal Unesp: Universidade Estadual Paulista 7 1. INTRODUÇÃO O progresso da medicina terapêutica só se tornou possível a partir do uso de modelos animais para se estudar a etiopatogenia e os mecanismos fisiopatológicos das doenças. Existem diversas semelhanças anatômicas, fisiológicas e genômicas entre humanos e suínos (Wernersson et al., 2005), fazendo com que essa espécie seja o modelo de eleição para estudos dos sistemas cardiovascular (Gabisonia et al., 2019; Hobby et al., 2019; Yuan et al., 2020), esquelético (Peck et al., 2015; Hai et al., 2017), urinário (Rodríguez et al., 2020) e de doenças metabólicas, neurodegenerativas e genéticas (Jakobsen et al, 2016, Yu et al., 2016; Cho et al., 2018). Em comparação com primatas, os suínos têm intervalos entre geração curtos, taxa de crescimento rápida, ninhada prolífica e técnicas de reprodução padronizadas (Lunney et al., 2021). A saúde do animal utilizado em estudos biomédicos é de responsabilidade do pesquisador e é crítica para a obtenção de resultados confiáveis (Kells, 2022). Suínos são pouco tolerantes à contenção física, o que pode causar alterações metabólicas, como hipertermia e dificuldade respiratória (Benjamin, 2005; Romero et al., 2016; Flores-Peinado, 2020). A contenção química (CQ), por sua vez, além de minimizar o estresse, pode ser associada à anestesia local para a realização de diversos procedimentos cirúrgicos, além de apresentar certas vantagens frente à anestesia geral, como: facilidade de administração; baixo custo e requerimento de equipamentos; e menor risco de complicações (Lin, 2022). A administração por via intramuscular (IM) tem sido amplamente utilizada na espécie suína para a administração de sedativos e tranquilizantes. No entanto, há o risco potencial de danos musculares e tempo de recuperação longo por deposição em tecido muscular (Kmiec, 2005; Mestorino et al., 2013; Hampton et al., 2021). Por outro lado, a administração intravenosa (IV) é um desafio em animais conscientes, pois o acesso venoso exige contenção física enérgica (Lin, 2022). 8 A administração intranasal (IN) é indolor e menos invasiva (Costantino et al., 2007) e a absorção dos fármacos é facilitada pelas vias neurais olfativas e trigeminais, sem que ocorra passagem pela barreira hematoencefálica (BHE) (Pardeshi e Belgamwar, 2013). O uso dessa via para administração de sedativos, tranquilizantes e analgésicos já foi descrito em várias espécies, incluindo humanos e suínos (Axiak et al., 2007; Becker et al., 2021; Hampton et al., 2021; Seak et al., 2021; Gómez-Manzano et al., 2022). Assim, buscou-se com este estudo propor uma alternativa para a contenção química de suínos, visando manter a homeostase ao minimizar o estresse da contenção. Para isso, foram comparadas as administrações intranasal e intramuscular da associação de azaperona, midazolam e cetamina (AMC) em suínos adultos, com relação às características da contenção química e à capacidade de manter a homeostase. Os objetivos específicos incluíram investigar se o período de latência e a duração da CQ após administração intranasal seriam menores do que na via intramuscular, analisar se a qualidade da CQ obtida pela administração intranasal seria comparável àquela alcançada pela via intramuscular, e examinar as alterações fisiológicas e possíveis efeitos adversos durante e após a CQ em ambas as vias de administração. 42 6. CONCLUSÃO A administração intranasal da associação AMC produziu contenção química imediata, com qualidade similar àquela obtida após a administração intramuscular, com mínimas alterações clínicas. 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