Delay classical eyeblink conditioning is an important model of associative, cerebellar dependent learning. Norepinephrine (NE) plays a significant modulatory role in the acquisition of learning; other neurotransmitter systems are also at play. The goal of this dissertation was to determine whether NE, GABA and glutamate (Glu) release is observed in cerebellar cortex during delay eye blink conditioning, and whether such release was selectively associated with training and not due only to stimulatory sensory input. The data support the hypothesis of noradrenergic and GABAergic system involvement in motor learning with NE as a modulator of early responding and GABA as a mediator of the learned response. In addition to neurotransmitter levels, we found that the local administration into the cerebellum of Rp-cAMP and propranolol impair the consolidation of learning when administered post training on the eyeblink conditioning task indicating that the B-adrenergic receptor and the cAMP downstream signaling cascade are essential for memory consolidation. These results support the hypothesis of NE acting as a neuromodulator in the cerebellum for the acquisition of motor learning. A similar experimental design was applied to aged animals and the neurochemical pattern of release was haracterized by a delay in the response to eyeblink conditioning and smaller amounts of the neurotransmitter evoked by the paired US-CS. It is hypothesized that the impairment in aging could be due to excitotoxicity caused by chronic inflammation. The present study also approached this issue by targeting the pro-inflammatory cytokine TNF-a and we found that suppression of TNF-a in aged animals improved learning.

Mesolimbic dopamine plays an important role in ethanol reinforcement, and studies have shown that accumbal dopamine increases during operant ethanol self-administration. However, no one has ever studied this dopaminergic response during the acquisition of ethanol self-administration. Furthermore, some studies have shown that the dopamine signal does not correlate with the pharmacological effects of ethanol, but with the time during which the animal consumes the majority of the ethanol solution and when the sensory stimuli of ethanol are strongest. However, there is currently no direct evidence showing that the sensory stimuli of ethanol is indeed what causes the brief increase in accumbal dopamine during ethanol self-administration. The studies in this dissertation attempted to elucidate these issues. We designed and tested a placebo spout, which was to be used to study the relationship between accumbal dopamine and the sensory stimuli of ethanol during self-administration. Unfortunately, the placebo designs were either not feasible for performing microdialysis or did not show promising behavioral data. We also developed and tested a self-administration protocol in which the concentrations of ethanol (10%) were kept constant throughout the study. The new protocol was successful in initiating and maintaining ethanol self-administration, and the animals doubled their intake from day 1 to day 2 of ethanol consumption. Using this protocol, we trained male Long Evans rats to self-administer ethanol and measured accumbal dopamine during the first two days of ethanol self-administration through microdialysis. The behavioral and neurochemical data matched. A single exposure to ethanol was sufficient for the animals to double their ethanol consumption by day 2 and to cause an increase in accumbal dopamine during the first 5 minutes of ethanol self-administration. The dopamine response was observed during the time when the sensory stimuli of ethanol were strongest, but before ethanol reached peak concentrations in the brain. Overall, these results suggest that the dopamine response to ethanol self-administration may not be solely pharmacological and that a single exposure to ethanol is sufficient to learn the association between ethanol and its cues. These findings give us greater insight into mesolimbic dopamine's role in the early stages of ethanol reinforcement. / text

The blood-brain barrier (BBB) controls the movement of substances into and out of the brain. The tight junctions between endothelial cells and energy dependent transporters in the BBB influence rate and extent of drug distribution to the brain. The aim of this thesis was to study different methodological and pharmacokinetic aspects of drug transport at the BBB by characterizing possible active uptake and drug-drug interactions. Therefore, advanced tools for data acquisition and analysis were applied. The role of BBB transport in early drug development, with particular emphasis on in vitro-in vivo comparisons and species differences, was also investigated. Microdialysis in rats was used to study the BBB pharmacokinetics of oxymorphone, diphenhydramine (DPHM), oxycodone and morphine. Oxymorphone, DPHM and verapamil were all found to be actively taken up at the BBB, with brain to blood unbound drug ratios of 2, 5 and 2, respectively. The effect profile for oxycodone was successfully described using the modified M3 method for censored observations. In vitro experiments indicated a competitive interaction between DPHM and oxycodone on active uptake transport to the brain. No such interaction was observed in vivo due to much lower unbound concentrations achieved, compared with the in vitro Ki values. Active uptake of morphine at the BBB was not demonstrated even at very low concentrations as it was not possible to separate the active uptake transport process from active efflux by decreasing the morphine concentration. Mice carrying the human P-gp gene (hMDR1) were used to evaluate possible species differences in P-gp function. Differences were evident between the hMDR1 and normal mice in BBB penetration of various P-gp substrates and in the effect of blockers on P-gp function. Quantitative measurements of P-gp expression levels at the BBB and a comparison with human data are crucial for the future use of the hMDR1 model. In conclusion, this thesis reports active uptake of oxymorphone, DPHM and verapamil at the BBB. In vivo interaction of DPHM and oxycodone at the BBB was found not to be significant at therapeutic drug concentrations. Furthermore species differences were found between human and mouse P-gp function at the BBB.

Advantages and disadvantages of administration of resuscitation fluids in burns patients have been discussed at length. However, the effect of resuscitation fluids on tissue physiological endpoints and tissue antibiotic distribution is scarcely reported, yet clinically crucial. The preliminary studies of this thesis involved evaluation of the literature and the development of a non - recovery anaesthetized rat model of burn injury suitable for the study of plasma and tissue physiological changes and antibiotic pharmacokinetics (PK). Therefore, the first series of the studies for this thesis was designed to examine the relative effects of a range of crystalloid and colloid-containing resuscitation fluids on tissue pH following burn injury in a rat model. The secondary aims were to examine the effects of these fluids on tissue blood flow, plasma protein extravasation (PPE) and evaporative water loss (EWL). In these studies we confirmed that the burn injury and fluid resuscitation were accompanied by a tissue acidosis. Administration of Lactated Ringers’ Albumin (LRA) and Lactated Ringers’ Dextran (LRD) effectively attenuated the degree of tissue acidosis in the thermally injured and non injured sites for 180 minutes post burn and the transepidermal water loss (TEWL) on the non injured sites during the first 60 minutes of the acute phase of burn injury. The second phase of the work was designed to assess the changes in antibiotic distribution with the administration of these different fluids in plasma as well as in interstitial tissues in the burn and the non burn sites. This study showed that for cephalothin (4g/kg body weight, administered intravenously (IV)), Lactated Ringers solution (LR) and Hypertonic Saline (HS) showed similar plasma PK with Time > Minimum inhibitory concentration (MIC) (> 180 minutes) in plasma. However, the antibiotic tissue distribution was more skewed towards lower levels for HS when compared with LR. For piperacillin (18g/kg body weight, administered IV), Time > MIC was considerably low comparatively, being only 55 min for both LR and HS. Antibiotic concentrations did not reach the MIC with LRA resuscitation. When considering the interstitial tissues, Time > MIC for cephalothin was lower than HS with LR on both the burn and the non burn sites. T > MIC for piperacillin was zero for all fluids in both burn and non burn sites. The major finding of this study was that with LRA resuscitation, antibiotic distribution was significantly lower than seen with LR and HS for both antibiotics studied in the interstitial tissue fluid space in both the burn and non burn sites. The final phase of the work was designed to study the apparent permeability co efficient of Keratinocytes (KC) to antibiotics in the presence of simulated pH changes observed in burn tissue in thermal injury using colloids and crystalloids. This study found that there was no significant difference between the basolateral and apical concentrations of antibiotics observed neither with the different pH values nor with time. However, there was definitely a significant difference in the apparent permeability of the cells with LR vs LRA and that the permeability was higher with LR than LRA. This study confirmed that the presence of LR allows greater permeation of the antibiotic into the KC, and also that with LRA resuscitation, the antibiotic tends to stay at higher concentrations in the interstitial compartment. These studies demonstrate that choice of resuscitation fluid following burn injury can affect both changes in tissue physiology and antibiotic distribution, warranting further study in both animal models and patient populations.

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 6 uppsatser.

Thesis (M.S.)--University of Florida, 2004. / Typescript. Title from title page of source document. Document formatted into pages; contains 152 pages. Includes Vita. Includes bibliographical references.

Objetivo: O objetivo do presente estudo foi descrever a relação entre as concentrações plasmáticas totais e livres tumorais do etoposídeo (ETO) e a inibição do crescimento do tumor observada em ratos Wistar portadores de tumor Walker- 256 (W256) utilizando a modelagem farmacocinética/farmacodinâmica (PK/PD). Métodos: Os procedimentos com animais foram aprovados no CEUA/UFRGS sob o número 22302. Os experimentos de farmacocinética foram realizados para determinar concentrações plasmáticas e livres em duas regiões do tumor sólido W256 através de microdiálise. Após a administração do ETO nas doses de 10 ou 20 mg/kg i.v. bolus em ratos Wistar portadores de tumor W256, amostras de sangue e microdialisado de tecido do centro e periferia do tumor foram coletadas simultaneamente, até 7 h pós-dose, para determinar o fator de penetração no tumor. Um método analítico por CLAE-UV foi desenvolvido e validado para quantificação do etoposídeo nas amostras de plasma e dialisado. Os experimentos de farmacodinâmica foram conduzidos em ratos portadores de tumor W256 que receberam ETO 5 e 10 mg/kg i.v. bolus uma vez ao dia por 8 e 4 dias, respectivamente. O volume dos tumores foram monitorados diariamente durante 30 dias. Análise não-compartimental dos dados de PK foi realizada no WinNonlin®. A modelagem dos dados PK e PK/PD foi realizada no Monolix®, utilizando abordagem populacional. Os dados PK/PD foram analisados usando o modelo Simeoni TGI modificado através da introdução de uma função Emax para descrever a relação nãolinear entre a concentração plasmática e tumoral e o efeito. Resultados e Discussão: O método por CLAE-UV foi desenvolvido e validado para quantificar as amostras de ETO em plasma e tecido. A penetração do ETO no tumor foi maior na periferia (61 ± 15 % e 61 ± 29 %) do que no centro do tumor (34 ± 6 % e 28 ± 11 %) após administração das doses 10 e 20 mg/kg, respectivamente (ANOVA, α = 0.05). Um modelo de 4 compartimentos compreendendo uma distribuição saturável (cinética de Michaelis-Menten) nos compartimentos tumorais a partir do compartimento central modelou simultaneamente os perfis de concentração-tempo do ETO em plasma e em ambas regiões do tumor. O modelo populacional PK/PD Simeoni TGI–Emax foi capaz de descrever o efeito antitumoral dependente do regime de administração do ETO utilizando concentrações totais plasmáticas ou livres no tumor, resultando em um maior k2max (potência máxima) para as concentrações livres (25,8 mL.μg-1.dia-1 - intratumoral vs. 12,6 mL.μg-1.dia-1 - plasma total). Conclusões: Os resultados mostram que a utilização das concentrações livres do fármaco no tumor para a modelagem PK/PD pode fornecer um melhor entendimento da relação farmacocinética e farmacodinâmica e melhoram a capacidade de previsão do modelo, considerando que a eficácia dos fármacos antineoplásicos no tratamento de tumores sólidos é dependente da capacidade do fármaco em se distribuir no tecido tumoral. / Objective: The aim of this study was to describe the relationship between total plasma and free interstitial tumor etoposide (ETO) concentrations and the drug tumor growth inhibition observed in a Walker-256 (W256) tumor-bearing Wistar rat model using the pharmacokinetic/pharmacodynamic (PK/PD) modeling. Methods: The experiments with animals were approved by CEUA/UFRGS (protocol number 22302). Pharmacokinetic experiments were conducted to determine total plasma and free intratumoral concentrations in two regions of W256 solid tumor by microdialysis. After administration of ETO 10 or 20 mg/kg i.v. bolus to W256 tumorbearing Wistar rats, blood and tissue microdialysate samples from tumor center and periphery were simultaneously collected up to 7h to determine the tumor penetration factor. An analytical HPLC-UV method was developed and validated for quantification of ETO in plasma and microdialysate samples. The pharmacodynamic experiments were conducted in W256 tumor-bearing rats that received ETO 5 or 10 mg/kg i.v. bolus every day for 8 and 4 days, respectively. Tumor volumes were monitored daily for 30 days. Non-compartmental analysis of PK data was performed in WinNonlin®. The PK and PK/PD modeling by population approach were performed using Monolix®. PK/PD data were analyzed using a modification of Simeoni TGI model by introducing an Emax function to describe the nonlinear relationship between tumor and plasma concentrations and effect. Results and Discussion: The HLPCUV method was developed and validated to determine plasma and tissue samples of ETO. ETO tumor penetration was higher in the tumor periphery (61 ± 15 % and 61 ± 29 %) than center (34 ± 6 % and 28 ± 11 %) following 10 and 20 mg/kg doses, respectively (ANOVA, α = 0.05). A 4-compartment structural model comprising a saturable distribution (Michaelis-Menten kinetics) into the tumor compartments from the central compartment simultaneously described the ETO concentration–time profiles in plasma and both tumor regions. The PK/PD population Simeoni TGI–Emax model was capable of describing the schedule-dependent antitumor effects of ETO using total plasma or free tumor concentrations obtained in a W256-tumor bearing Wistar rat model, resulting in higher k2max (maximal potency) for free concentrations (25.8 mL.μg-1.day-1 - intratumoral vs. 12.6 mL.μg-1.day-1 total plasma). Conclusions: The results showed that the use of free intratumoral drug concentrations in the PK/PD modeling can provide a better understanding of the pharmacokinetics and pharmacodynamics relationship and improve the forecasting ability of the models considering that the efficacy of antineoplastic drugs in the treatment of solid tumors is dependent on the drug ability to distribute into the tumor.

OBJETIVO: Desenvolver e validar a técnica de CLAE-DE para o doseamento de dopamina (DA), ácido diidrofenilacético (DOPAC), ácido homovanílico (HVA) e 3-metoxitiramina (3-MT); validar a técnica de microdiálise (MD) cerebral em animais conscientes e avaliar o efeito do tratamento agudo com um extrato lipofílico das partes aéreas de Hypericum caprifoliatum (HCP) sobre os níveis cerebrais de DA e seus metabólitos. MATERIAIS E MÉTODOS: A validação analítica foi realizada conforme o preconizado pela ANVISA, com análise dos parâmetros linearidade, precisão, exatidão, especificidade e limite inferior de quantificação. A técnica de microdiálise foi realizada de forma clássica: guias de sonda foram implantadas nas regiões cerebrais de interesse por cirurgia estereotáxica; os dialisados foram obtidos, 48 horas após a cirurgia, através da perfusão da sonda com líquido cérebroespinhal artificial (1 μL/min) e coleta durante três horas, em intervalos de 20 min; as três primeiras coletas foram utilizadas para determinação da linha de base e só então foram administrados os tratamentos. Para a validação da MD a sonda foi implantada no estriado (A: -0,3; L: + 3,2; P: - 4,5) e o tratamento foi sulfato de anfetamina (2 mg/kg, s.c.). Para a avaliação do efeito de HCP, a sonda foi implantada no núcleo acumbens (A: +2,2; L: -1,5; P: -5,8) e o tratamento foi uma dose de 270 mg/kg do extrato (v.o.). RESULTADOS E DISCUSSÃO: Os parâmetros avaliados para validar o método analítico ficaram dentro dos limites especificados pela ANVISA. O sulfato de anfetamina produziu um aumento nos níveis extracelulares de DA em 160% e uma redução de DOPAC e de HVA em 80% e 50%, respectivamente. O extrato HCP não alterou o conteúdo intersticial de DA, DOPAC e HVA no núcleo acumbens. Os valores basais encontrados estão de acordo com dados relatados na literatura. Não foi possível quantificar 3-MT. CONCLUSÃO: A metodologia analítica e as condições experimentais de microdiálise utilizadas permitiram a mensuração de mudanças induzidas por anfetamina nos níveis extracelulares de DA e seus principais metabólitos demonstrando que os fundamentos da técnica estão estabelecidos em nossas condições. O regime de tratamento empregado com HCP não foi suficiente para alterar os níveis de DA e seus principais metabólitos no núcleo acumbens de ratos. / OBJECTIVE: The objectives of this work were to validate the HLPC-ED technique for measuring dopamine (DA), dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 3- methoxytyramine (3-MT); to put into operation the brain microdialysis technique (MD) in freely moving rats; to evaluate, by using MD, the effect of a single treatment with lipophilic extract from aerial parts of Hypericum caprifoliatum (HCP) on DA, DOPAC, HVA and 3-MT brain levels. MATERIALS AND METHODS: In order to validate the analytical technique we followed the parameters established by ANVISA for bioanalytical samples. The parameters were: precision, accuracy, specificity and also the lower limit of quantitation. The microdialysis technique was carried out following classical procedures: male Wistar rats were submitted to stereotaxic surgery for the guide cannula implantation in the brain regions of interest; 48 hours later, the microdiaysis probe was inserted and perfused with cerebrospinal fluid (1 μL/min). After baseline samples collecting (1 h), the treatments were administrated and samples were collected every 20 min, during 2h. To accomplish MD technique validation the animals were treated with amphetamine sulfate (2 mg/kg, s.c.) and the extracellular levels of DA, DOPAC e HVA were measured in striatum (A: -0,3; L: + 3,2; P: - 4,5). The effect of HCP acute treatment (270 mg/kg, p.o.) was evaluated in nucleus accumbens (A: +2,2; L: -1,5; P: -5,8). RESULTS AND DISCUSSION: The analytical parameters values found were within the acceptable ranges for bio-analytical limits specified by ANVISA. As expected, amphetamine sulfate administration significantly increased DA (160%) extracellular levels and decreased DOPAC (80%) and HVA (50%) levels. The quantification of 3-MT was not possible. Acute HCP administration did not affected DA, DOPAC and HVA levels in the nucleus accumbens. CONCLUSION: The HPLC-ED methodology and microdialysis procedures employed allowed the detection of amphetamineinduced changes in extracellular levels of DA and its metabolites demonstrating that the technical starting point was acceptable. The acute administration of 270 mg/kg, p.o. of HCP to rats was not sufficient to modify the extracellular levels of DA, DOPAC and HVA in the nucleus accumbens.

Objetivos: objetivo geral deste trabalho foi avaliar a penetração cutânea da isotretinoína nanoencapsulada e livre utilizando técnicas de microdiálise e tape stripping in vitro. Métodos: A viabilidade da utilização da técnica de microdiálise para avaliar o perfil farmacocinético da isotretinoína após aplicação tópica foi investigada através da determinação da recuperação relativa (RR) in vitro por diálise e retrodiálise. A influência da concentração, do fluxo de perfusão e a ligação do fármaco à tubulação das sondas de microdiálise foram investigadas. A metodologia analítica para quantificação do fármaco em microdialisado foi validada. Em seguida, as penetrações cutâneas da isotretinoína livre e nanoencapsulada incorporada em géis hidrofílicos foram comparadas através da técnica de tape stripping in vitro em células de difusão de Franz utilizando pele humana e pele de porco. Para garantir a integridade das formulações, a estabilidade físico-química das mesmas foi avaliada. Os resultados de penetração cutânea foram comparados com os resultados in vivo obtidos em trabalho prévio do grupo de pesquisa. Resultados e Conclusões: Um método analítico simples e rápido para a determinação de isotretinoína em microdialisado foi validado de acordo com o FDA. A RR mostrou-se concentração independente e observou-se que há diferenças significativas entre as RR avaliadas pelos dois métodos utilizados, sendo a recuperação por retrodiálise 2,7 a 3,5 vezes superior que a obtida por diálise para os fluxos investigados. O fármaco aderiu às tubulações da sonda de microdiálise devido à sua lipofilicidade. Os hidrogéis de isotretinoína apresentaram estabilidade durante 2 meses de estocagem à 4 °C. Os experimentos de tape stripping in vitro mostraram que a isotretinoína não foi encontrada no compartimento receptor após 8 h, para ambas as formulações. A nanoencapsulação aumentou a penetração e prolongou a liberação da isotretinoína no estrato córneo de ambas as peles. A penetração cutânea em ambas as peles mostrou proporções similares para as duas formulações embora diferentes quantidades de fármaco tenham sido detectadas no estrato córneo. A pele de porco, mais permeável que a pele humana, é apropriada para prever a penetração cutânea da isotretinoína no estrato córneo humano in vitro (R = 0,79). O método in vitro não foi capaz de prever a penetração cutânea da isotretinoína in vivo. / Objectives: The aim of the present work was to assess the cutaneous penetration of isotretinoin free and loaded into polymeric nanocapsules using microdialysis and tape stripping in vitro. Methods: The feasibility of using microdialysis to determine the pharmacokinetic profile of isotretinoin after topical application was investigated through assessment of relative recovery (RR) in vitro by dialysis and retrodialysis. The influence of isotretinoin concentration, perfusion flow rate and drug binding to the probes were determined. The analytical method for quantification of microdialysate samples was validated. Furthermore the cutaneous penetration of isotretinoin free and loaded nanocapsules incorporated in hydrogel formulations were compared by tape stripping in vitro using Franz-type diffusion cells with excised human and pig skin. In order to ensure the integrity of the formulations used in this study, the chemical and physical stabilities were evaluated. The results of cutaneous penetration were compared with the results of tape stripping in vivo acquired in a previous study of our group. Results and Conclusions: A simple and rapid analytical method for quantification of isotretinoin in microdialysate samples was validated according to FDA. RR was concentration independent but method dependent under the conditions investigated being the retrodialysis recovery 3.5 to 2.7 times higher than the dialysis. Isotretinoin bound to the microdialysis tubing due to its high lipophilicity. The hydrogels showed storage stability for 2 months at 4 °C. In vitro tape stripping in human and pig skin showed that no isotretinoin reaches the receptor compartment for both formulations up to 8 h. Nanoencapsulation increased isotretinoin skin penetration for both stratum cornea and prolonged drug release. Similar proportion of cutaneous penetration for human and pig skin were observed although different amounts of drug were detected at the stratum corneum of both skin specimens. Pig skin, more permeable than human skin, is suitable for predicting cutaneous penetration of isotretinoin in humans in vitro (R = 0.79). The in vitro experiments were not suitable to reflect the in vivo results for percutaneous penetration of isotretinoin.

Objetivo: O objetivo do presente estudo foi descrever a relação entre as concentrações plasmáticas totais e livres tumorais do etoposídeo (ETO) e a inibição do crescimento do tumor observada em ratos Wistar portadores de tumor Walker- 256 (W256) utilizando a modelagem farmacocinética/farmacodinâmica (PK/PD). Métodos: Os procedimentos com animais foram aprovados no CEUA/UFRGS sob o número 22302. Os experimentos de farmacocinética foram realizados para determinar concentrações plasmáticas e livres em duas regiões do tumor sólido W256 através de microdiálise. Após a administração do ETO nas doses de 10 ou 20 mg/kg i.v. bolus em ratos Wistar portadores de tumor W256, amostras de sangue e microdialisado de tecido do centro e periferia do tumor foram coletadas simultaneamente, até 7 h pós-dose, para determinar o fator de penetração no tumor. Um método analítico por CLAE-UV foi desenvolvido e validado para quantificação do etoposídeo nas amostras de plasma e dialisado. Os experimentos de farmacodinâmica foram conduzidos em ratos portadores de tumor W256 que receberam ETO 5 e 10 mg/kg i.v. bolus uma vez ao dia por 8 e 4 dias, respectivamente. O volume dos tumores foram monitorados diariamente durante 30 dias. Análise não-compartimental dos dados de PK foi realizada no WinNonlin®. A modelagem dos dados PK e PK/PD foi realizada no Monolix®, utilizando abordagem populacional. Os dados PK/PD foram analisados usando o modelo Simeoni TGI modificado através da introdução de uma função Emax para descrever a relação nãolinear entre a concentração plasmática e tumoral e o efeito. Resultados e Discussão: O método por CLAE-UV foi desenvolvido e validado para quantificar as amostras de ETO em plasma e tecido. A penetração do ETO no tumor foi maior na periferia (61 ± 15 % e 61 ± 29 %) do que no centro do tumor (34 ± 6 % e 28 ± 11 %) após administração das doses 10 e 20 mg/kg, respectivamente (ANOVA, α = 0.05). Um modelo de 4 compartimentos compreendendo uma distribuição saturável (cinética de Michaelis-Menten) nos compartimentos tumorais a partir do compartimento central modelou simultaneamente os perfis de concentração-tempo do ETO em plasma e em ambas regiões do tumor. O modelo populacional PK/PD Simeoni TGI–Emax foi capaz de descrever o efeito antitumoral dependente do regime de administração do ETO utilizando concentrações totais plasmáticas ou livres no tumor, resultando em um maior k2max (potência máxima) para as concentrações livres (25,8 mL.μg-1.dia-1 - intratumoral vs. 12,6 mL.μg-1.dia-1 - plasma total). Conclusões: Os resultados mostram que a utilização das concentrações livres do fármaco no tumor para a modelagem PK/PD pode fornecer um melhor entendimento da relação farmacocinética e farmacodinâmica e melhoram a capacidade de previsão do modelo, considerando que a eficácia dos fármacos antineoplásicos no tratamento de tumores sólidos é dependente da capacidade do fármaco em se distribuir no tecido tumoral. / Objective: The aim of this study was to describe the relationship between total plasma and free interstitial tumor etoposide (ETO) concentrations and the drug tumor growth inhibition observed in a Walker-256 (W256) tumor-bearing Wistar rat model using the pharmacokinetic/pharmacodynamic (PK/PD) modeling. Methods: The experiments with animals were approved by CEUA/UFRGS (protocol number 22302). Pharmacokinetic experiments were conducted to determine total plasma and free intratumoral concentrations in two regions of W256 solid tumor by microdialysis. After administration of ETO 10 or 20 mg/kg i.v. bolus to W256 tumorbearing Wistar rats, blood and tissue microdialysate samples from tumor center and periphery were simultaneously collected up to 7h to determine the tumor penetration factor. An analytical HPLC-UV method was developed and validated for quantification of ETO in plasma and microdialysate samples. The pharmacodynamic experiments were conducted in W256 tumor-bearing rats that received ETO 5 or 10 mg/kg i.v. bolus every day for 8 and 4 days, respectively. Tumor volumes were monitored daily for 30 days. Non-compartmental analysis of PK data was performed in WinNonlin®. The PK and PK/PD modeling by population approach were performed using Monolix®. PK/PD data were analyzed using a modification of Simeoni TGI model by introducing an Emax function to describe the nonlinear relationship between tumor and plasma concentrations and effect. Results and Discussion: The HLPCUV method was developed and validated to determine plasma and tissue samples of ETO. ETO tumor penetration was higher in the tumor periphery (61 ± 15 % and 61 ± 29 %) than center (34 ± 6 % and 28 ± 11 %) following 10 and 20 mg/kg doses, respectively (ANOVA, α = 0.05). A 4-compartment structural model comprising a saturable distribution (Michaelis-Menten kinetics) into the tumor compartments from the central compartment simultaneously described the ETO concentration–time profiles in plasma and both tumor regions. The PK/PD population Simeoni TGI–Emax model was capable of describing the schedule-dependent antitumor effects of ETO using total plasma or free tumor concentrations obtained in a W256-tumor bearing Wistar rat model, resulting in higher k2max (maximal potency) for free concentrations (25.8 mL.μg-1.day-1 - intratumoral vs. 12.6 mL.μg-1.day-1 total plasma). Conclusions: The results showed that the use of free intratumoral drug concentrations in the PK/PD modeling can provide a better understanding of the pharmacokinetics and pharmacodynamics relationship and improve the forecasting ability of the models considering that the efficacy of antineoplastic drugs in the treatment of solid tumors is dependent on the drug ability to distribute into the tumor.