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The aim of this study was to characterize the semen of the Peruvian Hairless Dog. Fifteen males, aged 1 to 5 years, registered by the Kennel Club Peruano were used to collect semen (one ejaculated per male) by digital manipulation. A basic spermatogram was performed and macroscopic (volume and colour) and microscopic (sperm motility and concentration, using light field microscopy) characteristics were recorded. Additionally, sperm function tests were performed: sperm viability (using SYBR-14 and propidium iodide), mitochondrial membrane potential (MMP, using MitoTraker Deep Red FM), and acrosomal integrity (using FITC-PSA and propidium iodide) and analysed by flow cytometry with image analysis. The following values ​​were found (mean ± 95% CI): 6.1 ± 2.7 ml volume, opalescent colour; 66.7 ± 10.5% motility; 262.7 ± 20.4 x 106 sperm/ml sperm concentration; 77.7 ± 9.7% of sperm viability; 85.9 ± 5.1% high MMP; 83.6 ± 14.9% acrosomal integrity. These results represent the first report on the seminal characteristics of this breed and could be taken as reference values ​​for a better selection of breeding males and for the diagnosis of reproductive alterations of the breed., El objetivo del estudio fue caracterizar el semen del Perro sin Pelo del Perú. Quince machos, de edades entre 1 a 5 años, inscritos en el Kennel Club Peruano fueron utilizados para colectar semen (un eyaculado por macho) mediante manipulación digital. Se realizó un espermatograma básico y se registraron características macroscópicas (volumen y color) y microscópicas (motilidad y concentración espermática, utilizando microscopía de campo claro). Adicionalmente, se realizaron pruebas de función espermática: viabilidad espermática (utilizando SYBR-14 y ioduro de propidio), potencial de membrana mitocondrial (PMM; utilizando MitoTraker Deep Red FM), e integridad acrosomal (utilizando FITC-PSA y ioduro de propidio), analizadas posteriormente mediante citometría de flujo con análisis de imágenes. Se encontraron los siguientes valores (medias ± IC 95%): 6.1 ± 2.7 ml de volumen, color blanquecino; 66.7 ± 10.5% de motilidad; 262.7 ± 20.4 x 106 espermatozoides/ml de concentración espermática; 77.7 ± 9.7% de viabilidad espermática; 85.9 ± 5.1% de alto PMM; 83.6 ± 14.9% de integridad acrosomal. Estos resultados representan el primer reporte sobre las características seminales de esta raza y podrían ser tomados como valores de referencia para una mejor selección de machos reproductores y para el diagnóstico de alteraciones reproductivas de la raza.

The objective of this study was to detect changes in intracellular reactive oxygen species (superoxide anion and hydrogen peroxide) production and lipid peroxidation during cryopreservation of alpaca spermatozoa. Twelve alpaca semen samples were conventionally cryopreserved. Intracellular superoxide anion and hydrogen peroxide were evaluated by fluorescence microscopy using dihydroethidium (DHE)/YO-PRO-1 and dichlorofluorescein diacetate (H2DCFDA)/propidium iodide (PI), respectively. Evaluations were performed during cooling curve at (1) 25°C, (2) 15°C, (3) 5°C/0 min, (4) 5°C/15 min, (5) 5°C/30 min and (6) after freezing/thawing. Evaluation of lipid peroxidation by measuring malondialdehyde (MDA) was performed at 25°C, 5°C/30 min and after thawing. Maximum percentages of total spermatozoa producing superoxide anion and hydrogen peroxide were found at 5°C/30 min (62.8 ± 6.3% and 30.5 ± 5.6%, respectively), and these results were higher (p .05) than initial (25°C: 10.8 ± 3.8% and 6.8 ± 0.7%, respectively) and after thawing (29.8 ± 9.5% and 7.5 ± 1.8%, respectively) values. However, considering only viable spermatozoa, production of superoxide anion and hydrogen peroxide during overall stabilization at 5°C (76% and91%, respectively) and after thawing (74.9 ± 5.0% and 78.9 ± 2.2%, respectively) was higher (p .05) than initial values at 25°C (38.7 ± 3.1% and 53.6 ± 2.0%, respectively). Lipid peroxidation at 25°C, 5°C/30 min, and post-thawing were 346.5 ± 99.8, 401.1 ± 64.8 and 527.7 ± 142.8 ng/ml MDA, respectively. These results showed that high percentage of viable alpaca spermatozoa produces intracellular reactive species oxygen (ROS) during the cryopreservation process of alpaca semen.

El objetivo del estudio fue caracterizar el semen del Perro sin Pelo del Perú. Quince machos, de edades entre 1 a 5 años, inscritos en el Kennel Club Peruano fueron utilizados para colectar semen (un eyaculado por macho) mediante manipulación digital. Se realizó un espermatograma básico y se registraron características macroscópicas (volumen y color) y microscópicas (motilidad y concentración espermática, utilizando microscopía de campo claro). Adicionalmente, se realizaron pruebas de función espermática: viabilidad espermática (utilizando SYBR-14 y ioduro de propidio), potencial de membrana mitocondrial (PMM; utilizando MitoTraker Deep Red FM), e integridad acrosomal (utilizando FITC-PSA y ioduro de propidio), analizadas posteriormente mediante citometría de flujo con análisis de imágenes. Se encontraron los siguientes valores (medias ± IC 95%): 6.1 ± 2.7 ml de volumen, color blanquecino; 66.7 ± 10.5% de motilidad; 262.7 ± 20.4 x 106 espermatozoides/ml de concentración espermática; 77.7 ± 9.7% de viabilidad espermática; 85.9 ± 5.1% de alto PMM; 83.6 ± 14.9% de integridad acrosomal. Estos resultados representan el primer reporte sobre las características seminales de esta raza y podrían ser tomados como valores de referencia para una mejor selección de machos reproductores y para el diagnóstico de alteraciones reproductivas de la raza.

Epididymal alpaca sperm represent an alternative model for the study of alpaca semen. The objective of this study was to characterize the normal values of some functional variables in epididymal alpaca sperm using imaging flow cytometry. Alpaca testicles (n=150) were processed and sperm were recovered from the cauda epididymides. Only 76 samples with acceptable motility and sperm count were considered for assessment by imaging flow cytometry. Acrosome integrity and integrity/viability were assessed by FITC-PSA/PI and FITC-PNA/PI. Mitochondrial membrane potential (MMP) was assessed by MitoTracker CMXRos and MitoTracker Deep Red FM. Lipid peroxidation was evaluated using BODIPY 581/591 C11. Results show that the mean values for acrosome-intact sperm were 95.03±6.39% and 93.34±7.96%, using FITC-PSA and FITC-PNA, respectively. The mean values for acrosome-intact viable sperm were 60.58±12.12% with FITC-PSA/PI and 58.81±12.94% with FITC-PNA/PI. Greater MMP was detected in 65.03±15.92% and 59.52±19.19%, using MitoTracker CMXRos and MitoTracker Deep Red FM, respectively. Lipid peroxidation was 0.84±0.95%. Evaluation of acrosome-intact and acrosome-intact viable sperm with FITC-PSA/PI compared with. FITC-PNA/PI or MMP with MitoTracker CMXRos compared with MitoTracker Deep Red FM were correlated (P0.05). The MMP using MitoTracker CMXRos was the only variable correlated (P0.05) with sperm motility (r=0.3979). This report provides a basis for future research related to alpaca semen using the epididymal sperm model.

The main objective was to study the effects, on sperm function, of the addition of two superoxide dismutase (SOD) analogues (Tempo and Tempol) to alpaca semen extender for cryopreservation. Twelve alpaca semen samples were collected using an artificial vagina and then diluted at a 1:3 ratio in an extender based on skim milk, egg yolk, and fructose. Each semen sample was divided into three equal parts to form the following groups: control, Tempo (1 mM), and Tempol (1 mM). Groups were cooled to 5 °C in 90 minutes (-1 °C in 3 minutes); when samples reached approximately 10 °C, SOD analogues were added to the respective groups. At 5 °C, ethylene glycol (final concentration, 0.1 M) was added to each group. After 30 minutes at 5 °C, samples were loaded in 0.25 mL plastic straws, placed in liquid nitrogen vapor for 15 minutes, and then plunged. Percentages of sperm motility, functional sperm membrane integrity, and viable sperm with intact acrosomes were evaluated before and after freeze-thaw using visual analysis, the hypoosmotic swelling test, and the double-stain trypan blue/giemsa technique, respectively. The Terminal deoxymucleotidyl transferase dUTP Nick End Labeling assay was performed for evaluation of sperm DNA fragmentation of frozen-thawed sperm. Sperm motility was higher (P < 0.05) in the Tempol and Tempo groups than in the control group (mean, 22.1%, 19.7%, and 11.2%, respectively), with similar results for functional sperm membrane integrity. Additionally, DNA fragmentation was lower (P < 0.05) in the Tempol group (16.7%) than in the control group (38.8%). Viable sperm with intact acrosomes were not affected by the use of SOD analogues. There was a negative correlation (r = -0.58) between DNA fragmentation of alpaca sperm and sperm motility after freeze-thawing, but DNA damage was neither related to functional membrane integrity nor viable sperm with intact acrosomes. We concluded that DNA fragmentation and loss of motility during cryopreservation of alpaca sperm could be partially prevented by supplementation of the semen extender with 1 mM Tempo or Tempol.

The objective was to evaluate the effect of different cryoprotectant agents in the cryopreservation of Peruvian Paso horse semen. Twenty semen samples were collected from five Peruvian Paso horse stallions. Each sample was divided into 12 parts to form the groups: dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), ethylene glycol (EG) and glycerol (GLY), at 3%, 4% and 5%. Samples were frozen using a rate-controlled freezer. Sperm parameters evaluated were motility and viability/acrosomal status. After thawing, progressive motility in DMA group was higher (p

2 Páginas La descarga de GnRH del hipotálamo ocurre en forma intermitente en el día y la noche, esta descarga de GnRH produce la liberación de LH aproximadamente 30 minutos después, actuando sobre las células de Leydig, que inician la producción de progesterona, gran parte de la cual es transformada en testosterona, la cual tiene vida corta (20 a 60 minutos) y es de secreción pulsátil (Senger, 2003). La evaluación de los niveles séricos de testosterona en alpacas adultas a lo largo del año indica la existencia de diferencias estadísticas entre época reproductiva y no reproductiva como: 1142.50 ± 108.27, 992.50 ± 14.90, 877.50 ± 74.84 y 2445.00 ± 694.82 pg/mL en los meses de marzo, junio, septiembre y diciembre, respectivamente (Sumar et al., 1990). Dosis repetidas de 200 ng de GnRH en toros Holstein causaron un incremento en la producción seminal y las concentraciones de LH y Testosterona (Miller y Amman, 1986), mientras que dosis repetidas de 50 µg de GnRH en carneros produjeron un incremento en la concentración de testosterona sérica y un incremento de la actividad sexual (Schambancher y Lunstra, 1977). Uno de los factores que causan la baja fertilidad en las alpacas es la baja calidad seminal que presentan los machos, por lo cual se planteó esta investigación, con la intención de mejorar la calidad seminal mediante la utilización de GnRH exógena y evaluar el efecto de esta hormona sobre el perfil sérico de testosterona. Los objetivos del presente trabajo fueron determinar el efecto de la GnRH sobre las características seminales en alpacas machos y determinar las concentraciones séricas de testosterona post aplicación de GnRH. INTRODUCCIÓN. MATERIALES Y MÉTODOS. RESULTADOS Y DISCUSIÓN. CONCLUSIONES. BIBLIOGRAFÍA

Se emplearon 32 muestras de semen procedentes de cuatro ovinos a fin de ser criopreservadas con la adición dos antioxidantes: Tempo (2,2,6,6 tetrametil-1-piperidiniloxil) y Tempol (4-hidroxi 2,2,6,6 tetrametil-1-piperidiniloxil), en concentraciones de 0.5, 1.0 y 2.5 mM, para evaluar el efecto postdescongelamiento que pudieran tener sobre la motilidad progresiva, la viabilidad e integridad acrosomal y la capacitación espermática prematura. En cada concentración de los dos antioxidantes y del grupo control se utilizó un dilutor en base a Tris, realizándose ocho repeticiones, cada una con cuatro muestras de semen. Los resultados obtenidos demuestran que la adición de Tempo a una concentración de 0.5 mM mejora significativamente la calidad del semen criopreservado en comparación con el grupo control (p, Thirty-two semen samples from four rams were frozed using two antioxidants, Tempo (2,2,6,6 tetramethyl-1-piperidinyloxyl) and Tempol (4-hidroxi 2,2,6,6 tetramethyl-1- piperidinyloxyl), each one in concentrations of 0.5, 1.0, and 2.5 mM, to evaluate the effects on post-thawing motility, viability, acrosomal integrity, and earlier sperm capacitation. Eight replications for each antioxidant concentration and control group, were done using four semen samples per replication. Semen samples were diluted on a Tris extender. Results showed that Tempo 0.5 mM improved frozen semen quality in comparison with control group (p

2 páginas La oxitocina es una hormona que regula la contractibilidad del túbulo seminífero para facilitar el transporte de espermatozoides, además es un modulador de la esteroidogénesis (Nicholson, 1996); Oxitocina administrada en toros causa un incremento de la concentración espermática (Berndtson e Igboeli, 1988). Las características seminales fueron extensamente evaluados en alpacas, presentando alta variabilidad y mostrando valores diferentes a otras especies domesticas (Bravo, 2002). Los niveles séricos de testosterona en alpacas varían entre épocas, así tenemos: 1142.50 pg/mL en marzo y 877.50 ng/mL en septiembre (Sumar et al., 1990). El objetivo fue determinar el efecto de la oxitocina, sobre las características seminales en alpacas y la concentración sérica de testosterona post aplicación. INTRODUCCIÓN. MATERIALES Y MÉTODOS. RESULTADOS Y DISCUSIÓN. CONCLUSIONES.BIBLIOGRAFÍA.

El objetivo del presente trabajo fue evaluar el efecto del momento de adición del glicerol durante el proceso de enfriamiento sobre la calidad espermática del semen canino. Se utilizaron 15 eyaculados de 4 perros adultos. Cada muestra de semen fue dividida en alícuotas para someterlas a tres métodos de adición de glicerol (T1: al inicio; T2: al inicio y final; T3: al final de la curva de enfriamiento). El semen se envasó en pajillas de 0.5 ml y se congeló en nitrógeno líquido. Se evaluó la motilidad progresiva y la integridad funcional de membrana posdescongelamiento. No se encontró diferencia estadística entre los tres métodos de adición del glicerol, pudiéndose simplificar el proceso de criopreservación al añadir todo el glicerol al inicio de la curva de enfriamiento., The objective of the study was to evaluate the timing of adding glycerol during thecooling process on sperm quality of canine semen. Fifteen ejaculates from 4 mature dogswere used. Each sample was divided in aliquots and three methods of adding glycerolwere used (T1: at the beginning; T2; at the beginning and at the end; T3: at the end of thecooling process). Semen samples were packed in 0.5 ml straws and frozen in liquid nitrogen.The post-thawing progressive motility and the functional integrity of membrane wereevaluated. None statistical difference was observed due to the three treatments, indicatingthat the process of cryopreservation can be simplified by adding all the glycerol at thebeginning of the cooling curve.

Thirty-two semen samples from four rams were frozed using two antioxidants, Tempo (2,2,6,6 tetramethyl-1-piperidinyloxyl) and Tempol (4-hidroxi 2,2,6,6 tetramethyl-1- piperidinyloxyl), each one in concentrations of 0.5, 1.0, and 2.5 mM, to evaluate the effects on post-thawing motility, viability, acrosomal integrity, and earlier sperm capacitation. Eight replications for each antioxidant concentration and control group, were done using four semen samples per replication. Semen samples were diluted on a Tris extender. Results showed that Tempo 0.5 mM improved frozen semen quality in comparison with control group (p

Aim: To evaluate two extenders and two cryoprotectant agents (CPA) for alpaca semen cryopreservation. Methods: Semen samples were obtained from four adult alpacas (Lama pacos) and frozen using extender I (TRIS, citrate, egg yolk and glucose) or extender II (skim milk, egg yolk and fructose), each containing either glycerol (G) or ethylene glycol (EG) as CPA. Consequently, four groups were formed: 1) extender I-G; 2) extender I-EG; 3) extender II-G; and 4) extender II-EG. Semen was diluted in a two-step process: for cooling to 5 °C (extenders without CPA), and for freezing (extenders with CPA). Viability and acrosome integrity were assessed using trypan blue and Giemsa stains. Results: When compared, the motility after thawing was higher (P < 0.05) in groups II-EG (20.0 % ± 6.7 %) and II-G (15.3 % ± 4.1 %) than that in groups I-G (4.0 % ± 1.1 %) and I-EG (1.0 % ± 1.4 %). Viable spermatozoa with intact acrosomes in groups II-EG (18.7 % ± 2.9 %) and II-G (12.7 % ± 5.9 %) were higher than that in groups I-G (5.7 % ± 1.5 %) and I-EG (4.0 % ± 1.0 %). Conclusion: The skim milk- and egg yolk-based extenders containing ethylene glycol or glycerol to freeze alpaca semen seems to promote the survival of more sperm cells with intact acrosomes than the other extenders. (Asian J Androl 2005 Sep; 7: 303–309)