2-Aryladenine Derivatives as a Potent Scaffold for Adenosine Receptor Antagonists: The 6-Morpholino Derivatives.

A set of 2-aryl-9-H or methyl-6-morpholinopurine derivatives were synthesized and assayed through radioligand binding tests at human A1, A2A, A2B, and A3 adenosine receptor subtypes. Eleven purines showed potent antagonism at A1, A3, dual A1/A2A, A1/A2B, or A1/A3 adenosine receptors. Additionally, three compounds showed high affinity without selectivity for any specific adenosine receptor. The structure-activity relationships were made for this group of new compounds. The 9-methylpurine derivatives were generally less potent but more selective, and the 9H-purine derivatives were more potent but less selective. These compounds can be an important source of new biochemical tools and/or pharmacological drugs.

A Robust and Efficient FRET-Based Assay for Cannabinoid Receptor Ligands Discovery.

The identification of new modulators for Cannabinoid Receptors (CBRs) has garnered significant attention in drug discovery over recent years, owing to their manifold pathophysiological implications. In the context of hit identification, the availability of robust and sensitive high-throughput screening assays is essential to enhance the likelihood of success. In this study, we present the development and validation of a Tag-lite® binding assay designed for screening hCB1/hCB2 binding, employing a dual fluorescent ligand, CELT-335. Representative ligands for CBRs, exhibiting diverse affinity and functional profiles, were utilized as reference compounds to validate the robustness and efficiency of the newly developed Tag-lite® binding assay protocol. The homogeneous format, coupled with the sensitivity and optimal performance of the fluorescent ligand CELT-335, establishes this assay as a viable and reliable method for screening in hit and lead identification campaigns.

High-throughput nephelometry methodology for qualitative determination of aqueous solubility of chemical libraries.

The purpose of the protocol reported in this work is the solubility profiling of large chemical libraries using nephelometry. This technique allows the qualitative classification of compounds as highly, moderately, or poorly water-soluble. The described methodology is not intended to yield quantitative solubility values of the studied compounds but can be used as a primary solubility assessment of large chemical libraries, to guide hit prioritization after High Throughput Screening (HTS) campaigns.

Effect of Bulky N-Dibenzofuranylmethyl Substitution on the 5-HT2 Receptor Affinity and Efficacy of a Psychedelic Phenethylamine.

The introduction of arylmethyl substituents on the amine nitrogen atom of phenethylamines and tryptamines often results in profound increases in their affinity and functional activity at 5-HT2 serotonin receptors. To probe the sensitivity of this effect to substantially larger N-substituents, ten derivatives of the well-characterized psychedelic phenethylamine 2C-B were prepared by appending different dibenzo[b,d]furylmethyl (DBFM) moieties to the basic nitrogen. The DBFM group attached to the amino group through its 1-, -2-, or 3-position decreased affinity and agonist activity at the 5-HT2A/2C receptors. Substitution through the 4-position usually favored affinity for all three 5-HT2 receptor subtypes with compound 5 exhibiting 10- and 40-fold higher affinities at the 5-HT2A and 5-HT2C receptors, respectively, but less than fourfold selectivity among the three receptor subtypes. Nevertheless, all were relatively weak partial 5-HT2AR agonists, mostly in the low micromolar range, but full or nearly full agonists at the 5-HT2C subtype as determined in a calcium mobilization assay. Molecular docking simulations suggested that the dibenzofuryl portion dives more deeply into the orthosteric binding site of the 5-HT2A than the 5-HT2C receptor, interacting with the Trp3366.48 toggle switch associated with its activation, while the phenylamine moiety lies close to the extracellular side of the receptor. In conclusion, a very bulky N-substituent on a phenethylamine 5-HT2 receptor agonist is tolerated and may increase affinity if its orientation is appropriate. However, the Gq protein-mediated potencies are generally low, with low efficacy (relative to 5-HT) at the 5-HT2A receptor, somewhat higher efficacy at the 5-HT2B subtype, and full or nearly full efficacy at the 5-HT2C subtype.

Exploring Biginelli-based scaffolds as A2B adenosine receptor antagonists: Unveiling novel structure-activity relationship trends, lead compounds, and potent colorectal anticancer agents.

Antagonists of the A2B adenosine receptor have recently emerged as targeted anticancer agents and immune checkpoint inhibitors within the realm of cancer immunotherapy. This study presents a comprehensive evaluation of novel Biginelli-assembled pyrimidine chemotypes, including mono-, bi-, and tricyclic derivatives, as A2BAR antagonists. We conducted a comprehensive examination of the adenosinergic profile (both binding and functional) of a large compound library consisting of 168 compounds. This approach unveiled original lead compounds and enabled the identification of novel structure-activity relationship (SAR) trends, which were supported by extensive computational studies, including quantum mechanical calculations and free energy perturbation (FEP) analysis. In total, 25 molecules showed attractive affinity (Ki < 100 nM) and outstanding selectivity for A2BAR. From these, five molecules corresponding to the new benzothiazole scaffold were below the Ki < 10 nM threshold, in addition to a novel dual A2A/A2B antagonist. The most potent compounds, and the dual antagonist, showed enantiospecific recognition in the A2BAR. Two A2BAR selective antagonists and the dual A2AAR/A2BAR antagonist reported in this study were assessed for their impact on colorectal cancer cell lines. The results revealed a significant and dose-dependent reduction in cell proliferation. Notably, the A2BAR antagonists exhibited remarkable specificity, as they did not impede the proliferation of non-tumoral cell lines. These findings support the efficacy and potential that A2BAR antagonists as valuable candidates for cancer therapy, but also that they can effectively complement strategies involving A2AAR antagonism in the context of immune checkpoint inhibition.

G protein-specific mechanisms in the serotonin 5-HT2A receptor regulate psychosis-related effects and memory deficits.

G protein-coupled receptors (GPCRs) are sophisticated signaling machines able to simultaneously elicit multiple intracellular signaling pathways upon activation. Complete (in)activation of all pathways can be counterproductive for specific therapeutic applications. This is the case for the serotonin 2 A receptor (5-HT2AR), a prominent target for the treatment of schizophrenia. In this study, we elucidate the complex 5-HT2AR coupling signature in response to different signaling probes, and its physiological consequences by combining computational modeling, in vitro and in vivo experiments with human postmortem brain studies. We show how chemical modification of the endogenous agonist serotonin dramatically impacts the G protein coupling profile of the 5-HT2AR and the associated behavioral responses. Importantly, among these responses, we demonstrate that memory deficits are regulated by Gαq protein activation, whereas psychosis-related behavior is modulated through Gαi1 stimulation. These findings emphasize the complexity of GPCR pharmacology and physiology and open the path to designing improved therapeutics for the treatment of stchizophrenia.

N-Adamantyl-1-alkyl-4-oxo-1,4-dihydroquinoline-3-carboxamide Derivatives as Fluorescent Probes to Detect Microglia Activation through the Imaging of Cannabinoid Receptor Subtype 2 (CB2R).

Cannabinoid receptor subtype 2 (CB2R) is emerging as a pivotal biomarker to identify the first steps of inflammation-based diseases such as cancer and neurodegeneration. There is an urgent need to find specific probes that may result in green and safe alternatives to the commonly used radiative technologies, to deepen the knowledge of the CB2R pathways impacting the onset of the above-mentioned pathologies. Therefore, based on one of the CB2R pharmacophores, we developed a class of fluorescent N-adamantyl-1-alkyl-4-oxo-1,4-dihydroquinoline-3-carboxamide derivatives spanning from the green to the near-infrared (NIR) regions of the light spectrum. Among the synthesized fluorescent ligands, the green-emitting compound 55 exhibited a favorable binding profile (strong CB2R affinity and high selectivity). Notably, this ligand demonstrated versatility as its use was validated in different experimental settings such as flow cytometry saturation, competitive fluorescence assays, and in vitro microglia cells mimicking inflammation states where CB2R are overexpressed.

Confronting the challenge: a regional perspective by the Latin American pediatric infectious diseases society (SLIPE) expert group on respiratory syncytial virus-tackling the burden of disease and implementing preventive solutions.

Respiratory syncytial virus (RSV) is the leading cause of acute lower respiratory infections in children around the world. The post-pandemic era has resulted in a notable increase in reported cases of RSV infections, co-circulation of other respiratory viruses, shifts in epidemiology, altered respiratory season timing, and increased healthcare demand. Low- and middle-income countries are responsible for the highest burden of RSV disease, contributing significantly to health expenses during respiratory seasons and RSV-associated mortality in children. Until recently, supportive measures were the only intervention to treat or prevent RSV-infection, since preventive strategies like palivizumab are limited for high-risk populations. Advances in new available strategies, such as long-acting monoclonal antibodies during the neonatal period and vaccination of pregnant women, are now a reality. As the Regional Expert Group of the Latin American Pediatric Infectious Diseases Society (SLIPE), we sought to evaluate the burden of RSV infection in Latin America and the Caribbean (LAC) region, analyze current strategies to prevent RSV infection in children, and provide recommendations for implementing new strategies for preventing RSV infection in children in LAC region.

Polymeric nanocapsules loaded with poly(I:C) and resiquimod to reprogram tumor-associated macrophages for the treatment of solid tumors.

Background: In the tumor microenvironment (TME), tumor-associated macrophages (TAMs) play a key immunosuppressive role that limits the ability of the immune system to fight cancer. Toll-like receptors (TLRs) ligands, such as poly(I:C) or resiquimod (R848) are able to reprogram TAMs towards M1-like antitumor effector cells. The objective of our work has been to develop and evaluate polymeric nanocapsules (NCs) loaded with poly(I:C)+R848, to improve drug stability and systemic toxicity, and evaluate their targeting and therapeutic activity towards TAMs in the TME of solid tumors. Methods: NCs were developed by the solvent displacement and layer-by-layer methodologies and characterized by dynamic light scattering and nanoparticle tracking analysis. Hyaluronic acid (HA) was chemically functionalized with mannose for the coating of the NCs to target TAMs. NCs loaded with TLR ligands were evaluated in vitro for toxicity and immunostimulatory activity by Alamar Blue, ELISA and flow cytometry, using primary human monocyte-derived macrophages. For in vivo experiments, the CMT167 lung cancer model and the MN/MCA1 fibrosarcoma model metastasizing to lungs were used; tumor-infiltrating leukocytes were evaluated by flow cytometry and multispectral immunophenotyping. Results: We have developed polymeric NCs loaded with poly(I:C)+R848. Among a series of 5 lead prototypes, protamine-NCs were selected based on their physicochemical properties (size, charge, stability) and in vitro characterization, showing good biocompatibility on primary macrophages and ability to stimulate their production of T-cell attracting chemokines (CXCL10, CCL5) and to induce M1-like macrophages cytotoxicity towards tumor cells. In mouse tumor models, the intratumoral injection of poly(I:C)+R848-protamine-NCs significantly prevented tumor growth and lung metastasis. In an orthotopic murine lung cancer model, the intravenous administration of poly(I:C)+R848-prot-NCs, coated with an additional layer of HA-mannose to improve TAM-targeting, resulted in good antitumoral efficacy with no apparent systemic toxicity. While no significant alterations were observed in T cell numbers (CD8, CD4 or Treg), TAM-reprogramming in treated mice was confirmed by the relative decrease of interstitial versus alveolar macrophages, having higher CD86 expression but lower CD206 and Arg1 expression in the same cells, in treated mice. Conclusion: Mannose-HA-protamine-NCs loaded with poly(I:C)+R848 successfully reprogram TAMs in vivo, and reduce tumor progression and metastasis spread in mouse tumors.

Stapling Amantadine to Melanostatin Neuropeptide: Discovery of Potent Positive Allosteric Modulators of the D2 Receptors.

This work describes the synthesis and pharmacological and toxicological evaluation of melanostatin (MIF-1) bioconjugates with amantadine (Am) via a peptide linkage. The data from the functional assays at human dopamine D2 receptors (hD2R) showed that bioconjugates 1 (EC50 = 26.39 ± 3.37 nM) and 2 (EC50 = 17.82 ± 4.24 nM) promote a 3.3- and 4.9-fold increase of dopamine potency, respectively, at 0.01 nM, with no effect on the efficacy (Emax = 100%). In this assay, MIF-1 was only active at the highest concentration tested (EC50 = 23.64 ± 6.73 nM, at 1 nM). Cytotoxicity assays in differentiated SH-SY5Y cells showed that both MIF-1 (94.09 ± 5.75%, p < 0.05) and carbamate derivative 2 (89.73 ± 4.95%, p < 0.0001) exhibited mild but statistical significant toxicity (assessed through the MTT reduction assay) at 200 µM, while conjugate 1 was found nontoxic at this concentration.

Estado actual de la poliomielitis en Latinoamérica

Resumen La vacuna oral contra el poliovirus (OPV) ha sido fundamental en controlar la epidemia de poliomielitis, y destaca por su seguridad, eficacia, facilidad de administración oral y bajo costo. Sin embargo, a pesar de estas ventajas, al tratarse de una vacuna con virus vivos atenuados, existe la posibilidad de mutaciones que confieran neurovirulencia. Por ende, es importante la vigilancia de parálisis flácida aguda (PFA), ya sea asociada a las vacunas atenuadas (VAPP) o a los virus derivados de vacunas (VDPV). En esta revisión presentamos datos importantes de Latinoamérica en los últimos años, donde se revisan los datos de VDPV de transmisión comunitaria, de origen ambiguo y asociadas con inmunodeficiencias. Debido a la presencia de VDPV, es importante fortalecer el sistema de vigilancia epidemiológica por PFA, con datos muy inferiores a los recomendados en estos últimos años en las Américas. Adicionalmente, es fundamental mejorar las coberturas vacunales para reducir la cantidad de lactantes en riesgo de adquirir poliomielitis. En consecuencia, presentamos las tasas de cobertura vacunal con la vacuna inactivada contra el poliovirus (IPV) en la región y analizamos los programas de vacunación contra la poliomielitis en concordancia con las recomendaciones de la Sociedad Latinoamericana de Infectología Pediátrica (SLIPE; mínimo 3 dosis de IPV) y del Grupo de Expertos en Asesoramiento Estratégico (SAGE) sobre Inmunización de la OMS (mínimo 2 dosis de IPV). El estudio concluye con recomendaciones de los autores para el cambio de OPV a uso exclusivo de IPV, para aumentar las coberturas vacunales y para reforzar la vigilancia por PFA en la región.

Abstract Oral poliovirus vaccine (OPV) has been instrumental in controlling the polio epidemic, and stands out for its safety, efficacy, ease of oral administration, and low cost. However, despite these advantages, as it is a live attenuated virus vaccine, there is the possibility of mutations that confer neurovirulence. Therefore, surveillance for acute flaccid paralysis (AFP) is important, whether associated with live vaccines (VAPP) or vaccine-derived viruses (VDPV). In this review we present important data from Latin America in recent years, where data on VDPV of community transmission, of ambiguous origin and associated with immunodeficiencies are reviewed. Due to the presence of VDPV, it is important to strengthen the epidemiological surveillance system for AFP, with data much lower than those recommended in recent years in the Americas. Additionally, it is essential to improve vaccination coverage to reduce the number of infants at risk of acquiring poliomyelitis. Consequently, we present the vaccination coverage rates with the inactivated vaccine against poliovirus (IPV) in the region and analyze the vaccination programs against poliomyelitis in accordance with the recommendations of the Latin American Society of Pediatric Infectious Diseases (SLIPE; minimum 3 doses of IPV) and the WHO Strategic Advisory Expert Group (SAGE) on Immunization (minimum 2 doses of IPV). The study concludes with recommendations from the authors for the change from OPV to exclusive use of IPV, to increase vaccination coverage and to strengthen surveillance for AFP in the region.

Valoración morfométrica de la cabeza del espermatozoide del cerdo doméstico según su edad / Morphometric evaluation of sperm head of domestic pig according their age

Se utilizó el Análisis Automatizado de la Morfometría Espermática (ASMA) con el fin de determinar las dimensiones de la cabeza del espermatozoide (DCE) en semen de cerdos domésticos según la edad, además de agrupar las medidas obtenidas en subpoblaciones espermáticas (SP). Se evaluaron 36 muestras de semen fresco y diluido de 20 cerdos los cuales se clasificaron en dos categorías. A: menores de 18 meses de edad y B: mayores de 18 meses de edad. Las DCE (Largo, µm/ Ancho, µm/ Área, µm 2 y Perímetro, µm) se analizaron en frotis teñidos con Hemacolor ® mediante Sperm-Class Analyser ® (SCA) y los valores obtenidos guardados en una base de datos. El procedimiento GLM fue utilizado para evaluar el efecto de la edad del cerdo sobre las DCE y el análisis de agrupamiento (FASTCLUS) para identificar las SP. Los espermatozoides provenientes de cerdos mayores de 18 meses de edad presentaron mayor longitud (8,84 vs. 8,95 µm) que los cerdos menores de 18 meses de edad, sin embargo, las medias correspondientes al ancho (4,44 vs. 4,32 µm), área (33,33 vs. 32,39µm 2) y perímetro (27,65 vs. 26,3 µm) fueron más pequeñas en los cerdos de mayor edad. Dos SP fueron obtenidas con el fin de ratificar las diferencias observadas entre las 2 categorías de edades evaluadas (P<0,001). La población que incluyó los espermatozoides con las mayores dimensiones disminuyó de 41,61 por ciento en cerdos menores de 18 meses a 20,78 por ciento en cerdos mayores de 18 meses. Contrariamente, la SP que contenía los espermatozoides de menor tamaño incrementó de un 58,39 por ciento en cerdos menores de 18 meses a 79,22 por ciento en cerdos mayores de 18 meses. En conclusión, la edad de los cerdos influye significativamente sobre las DCE. Los cerdos de mayor edad tienen 20 por ciento más de espermatozoides de menor tamaño que los cerdos más jóvenes.

Assisted Sperm Morphometry Analysis (ASMA) was used to determine the sperm head dimensions (DCE) of boar by age, and then the data set clustered in sperm subpopulations (SP). To this purpose were evaluated 36 fresh and diluted semen samples of 20 Dalland domestic pigs, which were classified in 2 categories: under 18 months old and over 18 months old. The DCE (Length, µm/ Width, µm/, Area, µm 2 / and Perimeter, µm) were analyzed in slides stained by Hemacolor ® by the Sperm-Class Analyser ® (SCA), and the mean measurements recorded. A GLM procedure was performed to evaluate the effects of boar age on sperm head dimensions and clustering analysis (FAST-CLUS procedure) to separate in SP. Spermatozoa collected from older boar (over 18 months old) had head length larger (8.84 vs. 8.95 µm) than younger boar (under 18 months old), however, the width (4.44 vs. 4.32 µm), area (33.33 vs. 32.39 µm 2) and perimeter (27.65 to 26.3 µm) were smaller in older boar than younger boar. Two SP were clustered in this trial toratify the differences between younger and older pigs. The mean values of each DCE among the SP were significantly dif-ferent (P<0.001). Thus, the percentage of representation of the subpopulation that includes those spermatozoa whose dimensions are the largest decreased from 41.61 percent in pigs under 18 months old to 20.78 percent in pigs over 18 months old. Whereas, the percent of representation of the SP containing the smallest sper-matozoa increased from 58.39 percent in pigs under 18 months old to 153 79.22 percent in pigs over 18 months old. In conclusion, the age of sexually mature domestic male pig had a significant effect on the morphometric traits of their spermatozoa. Older boar had 20 percent more of smaller spermatozoa than younger boar.

Caracterización morfométrica de la cabeza del espermatozoide porcino mediante análisis computarizado (resultados preliminares) / Morphometry characterization of boar sperm head with computer assited analysis (preliminary results)

Para determinar los parámetros morfométricos de la cabeza espermática en semen porcino, así como evidenciar la presencia de subpoblaciones espermáticas fueron evaluadas 20 muestras seminales de 10 verracos Dalland. Sobre semen fresco y refrigerado fue evaluada la motilidad, vitalidad, acrosomas alterados y/o ausentes y anormalidades espermáticas. Mediante el análisis automatizado de la morfología espermática (ASMA), en frotis teñidos con Hemacolor®, se realizaron las mediciones de la cabeza espermática: Longitud (µm), Ancho (µm), Área (µm2), Perímetro (µm) y función Largo/Ancho. El efecto del proceso de refrigeración sobre las variables de calidad seminal y morfometría, se analizaron utilizando el GLM (SAS®) y para identificar las subpoblaciones espermáticas, se utilizó el procedimiento FASTCLUS (SAS®). La refrigeración a 16°C por 24 horas no afectó las características de calidad seminal de los eyaculados, pero si afectó las características morfométricas. La longitud de la cabeza disminuyó de 8,82 a 8,71 mm, así como el perímetro de 30,08 a 29,05 µm, mientras que aumentaron los valores de ancho (4,36 a 4,45 µm) y área (33,13 a 33,14 µm2). Se identificaron tres subpoblaciones espermáticas, con valores de distribución de 28,45 por ciento para la subpoblación 1 (espermatozoides grandes), 51,20 por ciento para la subpoblación 2 (medianos) y 20,35 por ciento para la subpoblación 3 (pequeños), las cuales se ven alteradas significativamente durante el proceso de refrigeración a 16°C.

To determine the morphometric parameters of the sperm head, and identify the presence of separate sperm subpopulations in boar semen were evaluated 20 ejaculate samples of 10 boars. Sperm motility, viability, acrosome integrity and morphological abnormalities were evaluated on fresh and cooling semen samples. By means Assisted Sperm Morphometry Analysis (ASMA), in slides stained by Hemacolor®, were determined the morphometric dimensions: Length (µm), Width (µm), Area (µm2), Perimeter (µm), and function Length/Width. Effect of cooling procedure on variables of semen quality and morphometric parameters were analyzed using GLM (SAS®). For identify the sperm subpopulations was used FASTCLUS procedure (SAS®). Cooling at 16°C for 24 hours did not affect the parameters of semen quality, but affected morphometric characteristics. Sperm head length decreased of 8.82 to 8.71 µm, and the sperm head perimeter of 30.08 to 29.05 µm, however, the width (from 4.36 to 4.45 mm) and area sperm head increased (33.13 to 33.14 µm2). Our results demonstrated that three separate sperm subpopulations coexist in boar ejaculates, 28.45% in the subpopulation 1 (larges), 51.20% in the subpopulation 2 (average), and 20.35% in the subpopulation 3 (small). These sperm subpopulation changed their distribution during cooling process.

Head dimensions of Brahman and their crossbred bull spermatozoa are affected by cryopreservation

El objetivo de este estudio fue determinar el efecto de la criopreservación sobre las características morfométricas de las cabezas de espermatozoides de toros Brahman y sus mestizos. Cinco eyaculados fueron colectados de 4 toros y diluidos a 30°C en una solución de leche descremada-yema de huevo. Por cada muestra se hicieron dos frotis: uno del semen diluido, antes de su congelación en vapores de nitrógeno líquido, y otro de semen descongelado una semana después de la congelación. Todos los frotis fueron secados al aire y coloreados con Hemacolor®. Se analizaron las dimensiones de la cabeza espermática para un mínimo de 150 espermatozoides por muestra mediante el Sperm Class Analyser® (SCA). El procedimiento GLM se realizó para evaluar el efecto de la criopreservación sobre las dimensiones morfométricas de las cabezas espermáticas. Las cabezas espermáticas de los toros fueron significativamente (P<0,001) menores en los espermatozoides criopreservados que en las muestras frescas para la longitud (9,00 µm vs. 9,43 µm), el ancho (4,82 µm vs. 5,13 µm), el perímetro (32,46 µm vs. 33,69 µm) y el área (36,20 µm²vs. 39,97 µm²) para todos los toros. Así mismo, se encontraron diferencias (P<0,001) de todos los parámetros morfométricos de los toros evaluados, encontrándose dimensiones de cabeza menores en las muestras descongeladas. La variabilidad individual (CV) de las medidas de cabeza espermática de los toros osciló entre el 5,9 y el 10,2 por ciento para las muestras frescas y descongeladas, respectivamente. En conclusión, este estudio indica que el proceso de criopreservación de semen de toro afecta la morfometría, al reducir las dimensiones de la cabeza espermática de toros Brahman y sus cruces. Las diferencias entre los toros evaluados puede ser indicativo de diferencias individuales al proceso de criopreservación.

The objective of this study was to determine the effect of cryopreservation on morphometrics characteristic of Brahman and their crossbred bull sperm heads. Five ejaculates were collected from 4 bulls and diluted at 30°C in a skim milk-egg yolk extender. Two microscope slides were prepared from single extended sperm samples prior to freezing in nitrogen vapors, and another one after thawing, sperm smears were prepared as described above. All slides were air dried and stained with Hemacolor®. Sperm-head dimensions for a minimum of 150 sperm heads/samples were analysed from each sample by means of the Sperm-Class Analyser® (SCA), and the mean measurements recorded. A GLM procedure was performed to evaluate the effect of cryopreservation on sperm head morphometric dimensions. Bull sperm heads were significantly (P<0.001) smaller in frozen-thawed spermatozoa than in the extended samples for length (9.00 µm vs. 9.43 µm), width (4.82 µm vs. 5.13 µm), perimeter (32.46 µm vs. 33.69 µm) and area (36.20 µm2 vs. 39.97 µm2) for all bulls. Also, differences (P<0.001) were found within all bulls for whole morphometric parameters. The individual variability of sperm head measurements across all bulls ranged from 5.9% to 10.2% for fresh and thawed samples, respectively. In conclusion, the present study indicate that cryopreservation of bull semen did affect the morphometry to reduce the dimensions of Brahman and crossbred bull sperm heads. The differences among bulls may be indicative of the individual bull resistance to the cryopreservation process.