An investigation of the neural mechanisms of interval timing behaviour

Timing behaviour plays an important role in the daily living of individuals from a great variety of species. For example, organisms must be able to discriminate between the durations of relevant events (temporal discrimination) and to regulate their own behaviour in time (temporal differentiation). The processes that allow animals to adjust their behaviour to the temporal regularities of the environment have been studied using different procedures which model the relationship between time and behaviour. A taxonomy of timing based on the subject’s location in time with respect to the signalled duration has been proposed. When an organism judges the duration of an elapsed interval the timing is retrospective (e.g. interval bisection); when it responds during an elapsing duration the timing is immediate (e.g. fixed-interval peak procedure); and finally when it chooses between future delayed outcomes the timing is prospective (e.g. inter-temporal choice schedules). It has been proposed that the cortico-striato-thalamo-cortical (CSTC) circuits play a special role in interval timing and inter-temporal choice behaviour. This thesis examined whether performance of timing tasks by rats induces neuronal activation within the prefrontal cortex and corpus striatum, as revealed by Fos expression, and explored a new approach to analyzing performance in an inter-temporal choice schedule. Chapter 1 describes the literature which forms the background of the project. It reviews interval timing and inter-temporal choice methodology and theory, the neurobiological substrates underlying both kinds of behaviour, and finally Fos expression, as a marker of neuronal activation. Chapters 2-4 present experiments that examined whether, in intact rats, performance of different interval timing tasks was associated with neuronal activation in the dorsal striatum and prefrontal cortex, as revealed by expression of the Fos protein, the product of the immediate-early gene c-fos (Experiments 1-3). Chapters 5-7 present experiments focused on some behavioural and neurobiological aspects of inter-temporal choice behaviour. One purpose of these experiments was to develop an abbreviated approach to estimate the rate of delay discounting (K) and reinforcer size sensitivity parameter (Q) based on the Multiplicative Hyperbolic Model of inter-temporal choice (MHM), using the adjusting-delay schedule. Additionally a novel way of quantifying transitional behaviour in the adjusting-delay schedule was presented based on analysis of the power spectrum of cyclical changes in the adjusting delay, dB (Experiment 4). This approach was used to analyze data obtained from rats performing on the adjusting-delay schedule under methodological manipulations (Experiment 5) and neurobiological interventions (Experiment 6). Experiment 1 (Chapter 2) investigated whether, in intact rats, performance on the discrete-trials temporal discrimination task was associated with neuronal activation in the prefrontal cortex and corpus striatum, as revealed by enhanced Fos expression in these areas. Performance on temporal and light-intensity discrimination tasks was well described by a two-parameter logistic equation. The rats trained under the timing task showed increased Fos expression in the orbital prefrontal cortex (OPFC) and the nucleus accumbens (Acb) compared to the rats trained under the light-intensity discrimination task, indicating a substantial activation of these areas during the timing task. However, there was no evidence for involvement of the dorsal striatum in the performance of this task. Experiment 2 (Chapter 3) examined whether performance on an interval-bisection task in the range of milliseconds showed increased Fos expression in the prefrontal cortex and corpus striatum compared to performance under a light-intensity bisection task. Performance on both bisection tasks conformed to the conventional logistic psychometric function. The rats trained under the timing task showed increased Fos expression in the OPFC, infralimbic and prelimbic cortex and Acb compared to the rats trained under the light-intensity bisection task. The results provided no evidence for an involvement of the dorsal striatum in the performance of this task. Experiment 3 (Chapter 4) investigated whether performance on the fixed-interval peak procedure (FIPP) was associated with increased neuronal activity in the prefrontal cortex and corpus striatum, as revealed by Fos expression. The results showed that response rate during peak trials was characterized by a ‘Gaussian plus ramp’ function, with maximal responding (peak rate) occurring around the time of the reinforcement in the FI trials (peak time). Consistent with the results of Experiments 1 and 2, the concentration of Fos-positive neurones in the OPFC was greater in rats exposed to FIPP than in rats exposed to a VI schedule. However, the results did not provide any evidence for a specific involvement of the dorsal or ventral striatum in FIPP performance. In Experiment 4 (Chapter 5), rats made repeated choices on an adjusting-delay schedule. Indifference delays, calculated from adjusting delays in the last 10 sessions, were shorter when the sizes of reinforcers were 14 and 25 µl of a 0.6 M sucrose solution than when they were 25 and 100 µl of the same solution. The ratio of the indifference delays (d50) was significantly smaller than that predicted on the basis of an assumed linear relation between reinforcer size and instantaneous reinforcer value. Estimates of K and Q fell within the values reported previously. Adjusting delays in successive blocks of trials were analysed using the Fourier transform. The power spectra obtained from individual rats had a dominant frequency that corresponded to a period of oscillation of the adjusting delay between 30 and 100 trial blocks. Power in the dominant frequency band declined with extended training. Experiment 5 (Chapter 6) examined the pattern of oscillation of dB in an adjusting-delay schedule using the power spectrum analysis. The step-size in which the delay to the larger reinforcer (dB) increased or decreased was tested across two conditions. In Condition 1, dB increased or decreased (according to the rats’ choice) by 20% from block n to block n+1. In Condition 2, the step size was 10%. The power spectrum analysis showed that the period of oscillation of the dominant frequency of the spectrum was significantly longer under Condition 2 than under Condition 1. There was a consistent trend for the power of oscillation to be higher in the initial segment of the experiment in both conditions. Experiment 6 (Chapter 7) examined the effect of excitotoxic lesion of the core of the nucleus accumbens (AcbC) on K and Q in an adjusting delay schedule using the same protocol as Experiment 4. The effect of the lesion on the power spectrum parameters was also examined. The AcbC-lesioned group showed significantly lower values of d50 than the sham-lesioned group. The ratio of the indifference delays seen in both groups was substantially less than the value predicted on the basis of an assumed linear relation between reinforcer size and instantaneous reinforcer value. K was higher in the lesioned group than in the sham-lesioned group; Q was not affected by the AcbC lesion. Neither the spectral power within the dominant frequency band nor the period corresponding to the dominant frequency differed significantly between groups. The final chapter (Chapter 8) summarizes the findings of the experiments, and discusses their implications for the putative role of the prefrontal cortex, and ventral and dorsal striatum in interval timing and inter-temporal choice, and for theoretical models of these behaviours. The role of the dorsal striatum is questioned, while a possible role of the Acb in temporal processing is proposed. It is argued that an integrated model of interval timing and inter-temporal choice behaviour may require more than the processes of reinforcement and timing to account for both types of behaviour. Some possible directions for future research in this area are also discussed

An investigation of the neural mechanisms of interval timing behaviour

Timing behaviour plays an important role in the daily living of individuals from a great variety of species. For example, organisms must be able to discriminate between the durations of relevant events (temporal discrimination) and to regulate their own behaviour in time (temporal differentiation). The processes that allow animals to adjust their behaviour to the temporal regularities of the environment have been studied using different procedures which model the relationship between time and behaviour. A taxonomy of timing based on the subject’s location in time with respect to the signalled duration has been proposed. When an organism judges the duration of an elapsed interval the timing is retrospective (e.g. interval bisection); when it responds during an elapsing duration the timing is immediate (e.g. fixed-interval peak procedure); and finally when it chooses between future delayed outcomes the timing is prospective (e.g. inter-temporal choice schedules). It has been proposed that the cortico-striato-thalamo-cortical (CSTC) circuits play a special role in interval timing and inter-temporal choice behaviour. This thesis examined whether performance of timing tasks by rats induces neuronal activation within the prefrontal cortex and corpus striatum, as revealed by Fos expression, and explored a new approach to analyzing performance in an inter-temporal choice schedule. Chapter 1 describes the literature which forms the background of the project. It reviews interval timing and inter-temporal choice methodology and theory, the neurobiological substrates underlying both kinds of behaviour, and finally Fos expression, as a marker of neuronal activation. Chapters 2-4 present experiments that examined whether, in intact rats, performance of different interval timing tasks was associated with neuronal activation in the dorsal striatum and prefrontal cortex, as revealed by expression of the Fos protein, the product of the immediate-early gene c-fos (Experiments 1-3). Chapters 5-7 present experiments focused on some behavioural and neurobiological aspects of inter-temporal choice behaviour. One purpose of these experiments was to develop an abbreviated approach to estimate the rate of delay discounting (K) and reinforcer size sensitivity parameter (Q) based on the Multiplicative Hyperbolic Model of inter-temporal choice (MHM), using the adjusting-delay schedule. Additionally a novel way of quantifying transitional behaviour in the adjusting-delay schedule was presented based on analysis of the power spectrum of cyclical changes in the adjusting delay, dB (Experiment 4). This approach was used to analyze data obtained from rats performing on the adjusting-delay schedule under methodological manipulations (Experiment 5) and neurobiological interventions (Experiment 6). Experiment 1 (Chapter 2) investigated whether, in intact rats, performance on the discrete-trials temporal discrimination task was associated with neuronal activation in the prefrontal cortex and corpus striatum, as revealed by enhanced Fos expression in these areas. Performance on temporal and light-intensity discrimination tasks was well described by a two-parameter logistic equation. The rats trained under the timing task showed increased Fos expression in the orbital prefrontal cortex (OPFC) and the nucleus accumbens (Acb) compared to the rats trained under the light-intensity discrimination task, indicating a substantial activation of these areas during the timing task. However, there was no evidence for involvement of the dorsal striatum in the performance of this task. Experiment 2 (Chapter 3) examined whether performance on an interval-bisection task in the range of milliseconds showed increased Fos expression in the prefrontal cortex and corpus striatum compared to performance under a light-intensity bisection task. Performance on both bisection tasks conformed to the conventional logistic psychometric function. The rats trained under the timing task showed increased Fos expression in the OPFC, infralimbic and prelimbic cortex and Acb compared to the rats trained under the light-intensity bisection task. The results provided no evidence for an involvement of the dorsal striatum in the performance of this task. Experiment 3 (Chapter 4) investigated whether performance on the fixed-interval peak procedure (FIPP) was associated with increased neuronal activity in the prefrontal cortex and corpus striatum, as revealed by Fos expression. The results showed that response rate during peak trials was characterized by a ‘Gaussian plus ramp’ function, with maximal responding (peak rate) occurring around the time of the reinforcement in the FI trials (peak time). Consistent with the results of Experiments 1 and 2, the concentration of Fos-positive neurones in the OPFC was greater in rats exposed to FIPP than in rats exposed to a VI schedule. However, the results did not provide any evidence for a specific involvement of the dorsal or ventral striatum in FIPP performance. In Experiment 4 (Chapter 5), rats made repeated choices on an adjusting-delay schedule. Indifference delays, calculated from adjusting delays in the last 10 sessions, were shorter when the sizes of reinforcers were 14 and 25 µl of a 0.6 M sucrose solution than when they were 25 and 100 µl of the same solution. The ratio of the indifference delays (d50) was significantly smaller than that predicted on the basis of an assumed linear relation between reinforcer size and instantaneous reinforcer value. Estimates of K and Q fell within the values reported previously. Adjusting delays in successive blocks of trials were analysed using the Fourier transform. The power spectra obtained from individual rats had a dominant frequency that corresponded to a period of oscillation of the adjusting delay between 30 and 100 trial blocks. Power in the dominant frequency band declined with extended training. Experiment 5 (Chapter 6) examined the pattern of oscillation of dB in an adjusting-delay schedule using the power spectrum analysis. The step-size in which the delay to the larger reinforcer (dB) increased or decreased was tested across two conditions. In Condition 1, dB increased or decreased (according to the rats’ choice) by 20% from block n to block n+1. In Condition 2, the step size was 10%. The power spectrum analysis showed that the period of oscillation of the dominant frequency of the spectrum was significantly longer under Condition 2 than under Condition 1. There was a consistent trend for the power of oscillation to be higher in the initial segment of the experiment in both conditions. Experiment 6 (Chapter 7) examined the effect of excitotoxic lesion of the core of the nucleus accumbens (AcbC) on K and Q in an adjusting delay schedule using the same protocol as Experiment 4. The effect of the lesion on the power spectrum parameters was also examined. The AcbC-lesioned group showed significantly lower values of d50 than the sham-lesioned group. The ratio of the indifference delays seen in both groups was substantially less than the value predicted on the basis of an assumed linear relation between reinforcer size and instantaneous reinforcer value. K was higher in the lesioned group than in the sham-lesioned group; Q was not affected by the AcbC lesion. Neither the spectral power within the dominant frequency band nor the period corresponding to the dominant frequency differed significantly between groups. The final chapter (Chapter 8) summarizes the findings of the experiments, and discusses their implications for the putative role of the prefrontal cortex, and ventral and dorsal striatum in interval timing and inter-temporal choice, and for theoretical models of these behaviours. The role of the dorsal striatum is questioned, while a possible role of the Acb in temporal processing is proposed. It is argued that an integrated model of interval timing and inter-temporal choice behaviour may require more than the processes of reinforcement and timing to account for both types of behaviour. Some possible directions for future research in this area are also discussed

Morphology and distribution of the South American snake Chironius leucometapus (Serpentes: Colubridae)

Morfologia e distribuição da serpente sul-americana Chironius leucometapus (Serpentes: Colubridae). Chironius leucometapus foi descrita há 25 anos como uma subespécie de C. fuscus em uma área restrita do Departamento de Junín, no centro do Peru, sem mais espécimes relatados desde então. Dados de 17 novos espécimes revelam que C. leucometapus é uma espécie de ampla distribuição ao longo do sopé amazônico dos Andes, entre o centro do Peru e o norte do Equador. Novos dados morfológicos são apresentados, incluindo descamação, hemipênis, coloração em vida e a primeira descrição do crânio de uma espécie de Chironius com base em imagens de alta qualidade de tomografas computadorizadas. Elevamos para oito o número de espécies de Chironius para o Equador.Morfología y distribución de la serpiente sudamericana Chironius leucometapus (Serpentes: Colubridae). Chironius leucometapus fue descrita hace 25 años como una subespecie de C. fuscus en un área restringida del Departamento de Junín en el centro de Perú, sin que se hayan reportado más especímenes desde entonces. Datos de 17 especímenes nuevos revelan que C. leucometapus es una especie de amplia distribución a lo largo de las estribaciones amazónicas de los Andes entre Perú central y el norte de Ecuador. Se presentan datos morfológicos nuevos, que incluyen escamación, hemipenes, color en vida, y la primera descripción del cráneo de una especie de Chironius basada en imágenes de alta calidad de escaneados CT. El número de especies de Chironius en Ecuador es elevado a ocho.Morphology and distribution of the South American snake Chironius leucometapus (Serpentes: Colubridae). Chironius leucometapus was described more than 25 years ago as subspecies of C. fuscus from restricted area in the Department of Junín in central Peru, with no additional specimens reported since. Examination of 17 new specimens reveals that C. leucometapus is widespread along the Amazonian slopes of the Andes between central Peru and northern Ecuador. New morphological data including scalation, hemipenes, color in life, and the frst description of the skull of Chironius to be based on high-quality CT-scan images are presented. The number of species of Chironius from Ecuador is elevated to eight

Effect of orexin-B-saporin-induced lesions of the lateral hypothalamus on performance on a progressive ratio schedule

It has been suggested that a sub-population of orexinergic neurones whose somata lie in the lateral hypothalamic area (LHA) play an important role in regulating the reinforcing value of both food and drugs. This experiment examined the effect of disruption of orexinergic mechanisms in the LHA on performance on the progressive-ratio schedule of reinforcement, in which the response requirement increases progressively for successive reinforcers. The data were analysed using a mathematical model which yields a quantitative index of reinforcer value and dissociates effects of interventions on motor and motivational processes (Killeen, 1994). Rats were trained under a progressive-ratio schedule using food-pellet reinforcement. They received bilateral injections of conjugated orexin-B-saporin (OxSap) into the LHA or sham lesions. Training continued for a further 40 sessions after surgery. Equations were fitted to the response rate data from each rat, and the parameters of the model were derived for successive blocks of 10 sessions. The OxSap lesion reduced the number of orexin-containing neurones in the LHA by approximately 50% compared to the sham-lesioned group. The parameter expressing the incentive value of the reinforcer was not significantly altered by the lesion. However, the parameter related to the maximum response rate was significantly affected, suggesting that that motor capacity was diminished in the OxSap-lesioned group. The results indicate that OxSap lesions of the LHA disrupted food-reinforced responding on the progressive-ratio schedule. It is suggested that this disruption was brought about by a change in non-motivational (motor) processes

Activation of Serotonin 2C Receptors in Dopamine Neurons Inhibits Binge-like Eating in Mice

Acknowledgments and Disclosures This work was supported by the National Institutes of Health (Grant Nos. R01DK093587 and R01DK101379 [to YX], R01DK092605 to [QT], R01DK078056 [to MM]), the Klarman Family Foundation (to YX), the Naman Family Fund for Basic Research (to YX), Curtis Hankamer Basic Research Fund (to YX), American Diabetes Association (Grant Nos. 7-13-JF-61 [to QW] and 1-15-BS-184 [to QT]), American Heart Association postdoctoral fellowship (to PX), Wellcome Trust (Grant No. WT098012 [to LKH]), and Biotechnology and Biological Sciences Research Council (Grant No. BB/K001418/1 [to LKH]). The anxiety tests (e.g., open-field test, light-dark test, elevated plus maze test) were performed in the Mouse Neurobehavior Core, Baylor College of Medicine, which was supported by National Institutes of Health Grant No. P30HD024064. PX and YH were involved in experimental design and most of the procedures, data acquisition and analyses, and writing the manuscript. XC assisted in the electrophysiological recordings; LV-T assisted in the histology study; XY, KS, CW, YY, AH, LZ, and GS assisted in surgical procedures and production of study mice. MGM, QW, QT, and LKH were involved in study design and writing the manuscript. YX is the guarantor of this work and, as such, had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis. The authors report no biomedical financial interests or potential conflicts of interest.Peer reviewedPublisher PD

Regresión logística basado en género para registro de títulos de grado en Universidad Técnica Ambato

El propósito esencial de este trabajo investigativo fue establecer un modelo de regresión logística basado en el género para el análisis probabilístico adecuado de las variables que intervienen en el registro de títulos de grado de la Universidad Técnica de Ambato en la plataforma destinada por la Secretaría de Educación Superior, Ciencia, Tecnología e Innovación del Ecuador. Se utilizaron los datos de la base informática del sistema interno para el registro de titulaciones desarrollado por la Dirección de Tecnologías de la Información y Comunicación de la referida Institución. La población que fue objeto de estudio comprendió el periodo entre el 1 de julio de 2017 al 31 de julio de 2018. La presente investigación nace a partir de que los datos y variables correspondientes al registro de títulos de grado dentro de la Universidad mencionada, no reciben el tratamiento estadístico adecuado. Mediante la investigación de campo y a través de un enfoque cuantitativo de la información objeto de estudio, permitió determinar que la variable género influye o incide de manera directa sobre las principales variables que son inherentes a la graduación de un estudiante, y, que mediante una correcta determinación de un modelo de regresión logística en base al género, permite pronosticar o determinar el comportamiento de las variables pertinentes dentro del proceso de registro de título de un graduado, para el posible establecimiento de políticas o procesos que permitan estar a la vanguardia en lo relacionado a la equidad de géner

Anorectic and aversive effects of GLP-1 receptor agonism are mediated by brainstem cholecystokinin neurons, and modulated by GIP receptor activation

This work was funded by an MRC Career Development Award (MR/ P009824/1 and MR/P009824/2) to GD’A, as well as an MRC grant to SML/GD’A (MR/T032669/1), a BBSRC grant to SML (BB/M001067/1), and an additional direct contribution from Eli Lilly. D.J.H. was sup- ported by MRC (MR/N00275X/1 and MR/S025618/1), Diabetes UK (17/ 0005681), and the European Research Council (ERC) under the Eu- ropean Union’s Horizon 2020 research and innovation programme (Starting Grant 715884 to D.J.H.). AC was supported for part of this project by a travel grant from the Italian Society of Pharmacology and a fellowship from the Veronesi Foundation (Italy).Peer reviewedPublisher PD

Activation of Ventral Tegmental Area 5-HT2C Receptors Reduces Incentive Motivation

FUNDING AND DISCLOSURE The research was funded by Wellcome Trust (WT098012) to LKH; and National Institute of Health (DK056731) and the Marilyn H. Vincent Foundation to MGM. The University of Michigan Transgenic Core facility is partially supported by the NIH-funded University of Michigan Center for Gastrointestinal Research (DK034933). The remaining authors declare no conflict of interest. ACKNOWLEDGMENTS We thank Dr Celine Cansell, Ms Raffaella Chianese and the staff of the Medical Research Facility for technical assistance. We thank Dr Vladimir Orduña for the scientific advice and technical assistance.Peer reviewedPublisher PD

Lorcaserin improves glycemic control via a melanocortin neurocircuit.

OBJECTIVE: The increasing prevalence of type 2 diabetes (T2D) and associated morbidity and mortality emphasizes the need for a more complete understanding of the mechanisms mediating glucose homeostasis to accelerate the identification of new medications. Recent reports indicate that the obesity medication lorcaserin, a 5-hydroxytryptamine (5-HT, serotonin) 2C receptor (5-HT2CR) agonist, improves glycemic control in association with weight loss in obese patients with T2D. Here we evaluate whether lorcaserin has an effect on glycemia without body weight loss and how this effect is achieved. METHODS: Murine models of common and genetic T2D were utilized to probe the direct effect of lorcaserin on glycemic control. RESULTS: Lorcaserin dose-dependently improves glycemic control in mouse models of T2D in the absence of reductions in food intake or body weight. Examining the mechanism of this effect, we reveal a necessary and sufficient neurochemical mediator of lorcaserin's glucoregulatory effects, brain pro-opiomelanocortin (POMC) peptides. To clarify further lorcaserin's therapeutic brain circuit, we examined the receptor target of POMC peptides. We demonstrate that lorcaserin requires functional melanocortin4 receptors on cholinergic preganglionic neurons (MC4RChAT) to exert its effects on glucose homeostasis. In contrast, MC4RChAT signaling did not impact lorcaserin's effects on feeding, indicating a divergence in the neurocircuitry underpinning lorcaserin's therapeutic glycemic and anorectic effects. Hyperinsulinemic-euglycemic clamp studies reveal that lorcaserin reduces hepatic glucose production, increases glucose disposal and improves insulin sensitivity. CONCLUSIONS: These data suggest that lorcaserin's action within the brain represents a mechanistically novel treatment for T2D: findings of significance to a prevalent global disease

Molecular identification and frequency of isolated pathogens from bovine mastitis in dairy herds from Baja California Peninsula, Mexico

La mastitis bovina es una enfermedad de alto impacto económico para la industria lechera, y algunos de los agentes etiológicos que la provocan también son de interés en el ámbito de salud pública. El objetivo de este estudio fue identificar las especies bacterianas aisladas de casos de mastitis bovina, provenientes de siete establos lecheros ubicados en la Península de Baja California. Se tomaron 316 muestras de leche de igual número de cuartos, pertenecientes a 186 vacas en producción que a la prueba de California tuvieron reacción positiva. Se obtuvieron 182 aislados bacterianos de 163 cuartos pertenecientes a 106 vacas y se identificaron por PCR y secuenciación, dando un total de 20 especies diferentes. Además, se obtuvieron las frecuencias relativas, siendo los agentes causales más frecuentes: Staphylococcus aureus (58.8 %), Streptococcus agalactiae (13.2 %), Staphylococcus chromogenes (8.8 %), Escherichia coli (2.2 %) y Streptococcus uberis (2.2 %). El 6.13 % (10/163) de los cuartos con aislamiento presentaron infección mixta, siendo la combinación más frecuente S. aureus con S. agalactiae 30 % (3/10). Estos resultados indican una alta frecuencia y diversidad de patógenos de carácter contagioso y ambiental que provocan mastitis en ganado lechero en la región de estudio, siendo algunos de importancia para la salud pública. Los resultados observados, muestran que las causas de la mastitis son diversas, por lo que es indispensable mejorar las medidas de control y prevención, pero también establecer el diagnóstico de rutina para lograr controlar la mastitis.Bovine mastitis is a disease of high economic impact for the dairy industry and some of the etiological agents that cause it are also of interest in the field of public health. The purpose of study was to identify the bacterial causes of bovine mastitis from seven dairy farms located in the Baja California Peninsula. A total of 316 milk samples were collected from the same number of quarters belonging to 186 cows in production that tested positive for California Mastitis test. It was obtained 182 bacterial isolates from 163 quarters belonging to 106 cows and were identified by PCR, giving a total of 20 different species. Isolates were identified using specific oligonucleotides for the major mastitis pathogens and with universal oligonucleotides for the 16S ribosomal DNA gene with subsequent sequencing for those that did not amplify with the specific oligonucleotides and relative frequencies were obtained. The most frequent causal agents were: Staphylococcus aureus (58.8 %), Streptococcus agalactiae (13.2 %), Staphylococcus chromogenes (8.8 %), Escherichia coli (2.2 %) and Streptococcus uberis (2.2 %). A mixed infection was found in 6.13 % (10/163) of the quarters, being the most frequent combination S. aureus plus S. agalactiae 30 % (3/10). These results indicate a high frequency and diversity of contagious and environmental pathogens causing mastitis in dairy cattle in the region of study, being some of importance for public health. The results show that the causes of mastitis are diverse, so it is essential to improve control and prevention measures, but also to establish a routine diagnosis to control mastitis