Hypoxic Regulation of the Large-Conductance, Calcium and Voltage-Activated Potassium Channel, BK

Hypoxia is a condition characterized by a reduction of cellular oxygen levels derived from alterations in oxygen balance. Hypoxic events trigger changes in cell-signaling cascades, oxidative stress, activation of pro-inflammatory molecules, and growth factors, influencing the activity of various ion channel families and leading to diverse cardiovascular diseases such as myocardial infarction, ischemic stroke, and hypertension. The large-conductance, calcium and voltage-activated potassium channel (BK) has a central role in the mechanism of oxygen (O2) sensing and its activity has been related to the hypoxic response. BK channels are ubiquitously expressed, and they are composed by the pore-forming α subunit and the regulatory subunits β (β1–β4), γ (γ1–γ4), and LINGO1. The modification of biophysical properties of BK channels by β subunits underly a myriad of physiological function of these proteins. Hypoxia induces tissue-specific modifications of BK channel α and β subunits expression. Moreover, hypoxia modifies channel activation kinetics and voltage and/or calcium dependence. The reported effects on the BK channel properties are associated with events such as the increase of reactive oxygen species (ROS) production, increases of intracellular Calcium ([Ca2+]i), the regulation by Hypoxia-inducible factor 1α (HIF-1α), and the interaction with hemeproteins. Bronchial asthma, chronic obstructive pulmonary diseases (COPD), and obstructive sleep apnea (OSA), among others, can provoke hypoxia. Untreated OSA patients showed a decrease in BK-β1 subunit mRNA levels and high arterial tension. Treatment with continuous positive airway pressure (CPAP) upregulated β1 subunit mRNA level, decreased arterial pressures, and improved endothelial function coupled with a reduction in morbidity and mortality associated with OSA. These reports suggest that the BK channel has a role in the response involved in hypoxia-associated hypertension derived from OSA. Thus, this review aims to describe the mechanisms involved in the BK channel activation after a hypoxic stimulus and their relationship with disorders like OSA. A deep understanding of the molecular mechanism involved in hypoxic response may help in the therapeutic approaches to treat the pathological processes associated with diseases involving cellular hypoxia.

Transcriptome analysis of the hypothalamus and pituitary of turkey hens with low and high egg production

Background High egg producing hens (HEPH) show increased hypothalamic and pituitary gene expression related to hypothalamo-pituitary-gonadal (HPG) axis stimulation as well as increased in vitro responsiveness to gonadotropin releasing hormone (GnRH) stimulation in the pituitary when compared to low egg producing hens (LEPH). Transcriptome analysis was performed on hypothalamus and pituitary samples from LEPH and HEPH to identify novel regulators of HPG axis function. Results In the hypothalamus and pituitary, 4644 differentially expressed genes (DEGs) were identified between LEPH and HEPH, with 2021 genes up-regulated in LEPH and 2623 genes up-regulated in HEPH. In LEPH, up-regulated genes showed enrichment of the hypothalamo-pituitary-thyroid (HPT) axis. Beta-estradiol was identified as an upstream regulator regardless of tissue. When LEPH and HEPH samples were compared, beta-estradiol was activated in HEPH in 3 of the 4 comparisons, which correlated to the number of beta-estradiol target genes up-regulated in HEPH. In in vitro pituitary cell cultures from LEPH and HEPH, thyroid hormone pretreatment negatively impacted gonadotropin subunit mRNA levels in cells from both LEPH and HEPH, with the effect being more prominent in HEPH cells. Additionally, the effect of estradiol pretreatment on gonadotropin subunit mRNA levels in HEPH cells was negative, whereas estradiol pretreatment increased gonadotropin subunit mRNA levels in LEPH cells. Conclusions Up-regulation of the HPT axis in LEPH and upstream beta-estradiol activation in HEPH may play a role in regulating HPG axis function, and ultimately ovulation rates. Thyroid hormone and estradiol pretreatment impacted gonadotropin mRNA levels following GnRH stimulation, with the inhibitory effects of thyroid hormone more detrimental in HEPH and estradiol stimulatory effects more prominent in LEPH. Responsiveness to thyroid hormone and estradiol may be due to desensitization to thyroid hormone and estradiol in LEPH and HEPH, respectively, due to up-regulation of the HPT axis in LEPH and of the HPG axis in HEPH. Further studies will be necessary to identify possible target gene desensitization mechanisms and elicit the regulatory role of the HPT axis and beta-estradiol on ovulation rates in turkey hens.

Transcriptome analysis of the hypothalamus and pituitary of turkey hens with low and high egg production

Background: High egg producing hens (HEPH) show increased hypothalamic and pituitary gene expression related to hypothalamo-pituitary-gonadal (HPG) axis stimulation as well as increased in vitro responsiveness to gonadotropin releasing hormone (GnRH) stimulation in the pituitary when compared to low egg producing hens (LEPH). Transcriptome analysis was performed on hypothalamus and pituitary samples from LEPH and HEPH to identify novel regulators of HPG axis function. Results: In the hypothalamus and pituitary, 4644 differentially expressed genes (DEGs) were identified between LEPH and HEPH, with 2021 genes up-regulated in LEPH and 2623 genes up-regulated in HEPH. In LEPH, up-regulated genes showed enrichment of the hypothalamo-pituitary-thyroid (HPT) axis. Beta-estradiol was identified as an upstream regulator regardless of tissue. When LEPH and HEPH samples were compared, beta-estradiol was activated in HEPH in 3 of the 4 comparisons, which correlated to the number of beta-estradiol target genes up-regulated in HEPH. In in vitro pituitary cell cultures from LEPH and HEPH, thyroid hormone pretreatment negatively impacted gonadotropin subunit mRNA levels in cells from both LEPH and HEPH, with the effect being more prominent in HEPH cells. Additionally, the effect of estradiol pretreatment on gonadotropin subunit mRNA levels in HEPH cells was negative, whereas estradiol pretreatment increased gonadotropin subunit mRNA levels in LEPH cells.Conclusions: Up-regulation of the HPT axis in LEPH and upstream beta-estradiol activation in HEPH may play a role in regulating HPG axis function, and ultimately ovulation rates. Thyroid hormone and estradiol pretreatment impacted gonadotropin mRNA levels following GnRH stimulation, with the inhibitory effects of thyroid hormone more detrimental in HEPH and estradiol stimulatory effects more prominent in LEPH. Responsiveness to thyroid hormone and estradiol may be due to desensitization to thyroid hormone and estradiol in LEPH and HEPH, respectively, due to up-regulation of the HPT axis in LEPH and of the HPG axis in HEPH. Further studies will be necessary to identify possible target gene desensitization mechanisms and elicit the regulatory role of the HPT axis and beta-estradiol on ovulation rates in turkey hens.

Transcriptome analysis of the hypothalamus and pituitary of turkey hens with low and high egg production

Background: High egg producing hens ( HEPH ) show increased hypothalamic and pituitary gene expression related to hypothalamo-pituitary-gonadal ( HPG ) axis stimulation as well as increased in vitro responsiveness to gonadotropin releasing hormone ( GnRH ) stimulation in the pituitary when compared to low egg producing hens ( LEPH ). Transcriptome analysis was performed on hypothalamus and pituitary samples from LEPH and HEPH to identify novel regulators of HPG axis function. Results: In the hypothalamus and pituitary, 4644 differentially expressed genes ( DEGs ) were identified between LEPH and HEPH, with 2021 genes up-regulated in LEPH and 2623 genes up-regulated in HEPH. In LEPH, up-regulated genes showed enrichment of the hypothalamo-pituitary-thyroid ( HPT ) axis. Beta-estradiol was identified as an upstream regulator regardless of tissue. When LEPH and HEPH samples were compared, beta-estradiol was activated in HEPH in 3 of the 4 comparisons, which correlated to the number of beta-estradiol target genes up-regulated in HEPH. In in vitro pituitary cell cultures from LEPH and HEPH, thyroid hormone pretreatment negatively impacted gonadotropin subunit mRNA levels in cells from both LEPH and HEPH, with the effect being more prominent in HEPH cells. Additionally, the effect of estradiol pretreatment on gonadotropin subunit mRNA levels in HEPH cells was negative, whereas estradiol pretreatment increased gonadotropin subunit mRNA levels in LEPH cells. Conclusions: Up-regulation of the HPT axis in LEPH and upstream beta-estradiol activation in HEPH may play a role in regulating HPG axis function, and ultimately ovulation rates. Thyroid hormone and estradiol pretreatment impacted gonadotropin mRNA levels following GnRH stimulation, with the inhibitory effects of thyroid hormone more detrimental in HEPH and estradiol stimulatory effects more prominent in LEPH. Responsiveness to thyroid hormone and estradiol may be due to desensitization to thyroid hormone and estradiol in LEPH and HEPH, respectively, due to up-regulation of the HPT axis in LEPH and of the HPG axis in HEPH. Further studies will be necessary to identify possible target gene desensitization mechanisms and elicit the regulatory role of the HPT axis and beta-estradiol on ovulation rates in turkey hens.

Transcriptome analysis of the hypothalamus and pituitary of turkey hens with low and high egg production

Background: High egg producing hens ( HEPH ) show increased hypothalamic and pituitary gene expression related to hypothalamo-pituitary-gonadal ( HPG ) axis stimulation as well as increased in vitro responsiveness to gonadotropin releasing hormone ( GnRH ) stimulation in the pituitary when compared to low egg producing hens ( LEPH ). Transcriptome analysis was performed on hypothalamus and pituitary samples from LEPH and HEPH to identify novel regulators of HPG axis function. Results: In the hypothalamus and pituitary, 4644 differentially expressed genes ( DEGs ) were identified between LEPH and HEPH, with 2021 genes up-regulated in LEPH and 2623 genes up-regulated in HEPH. In LEPH, up-regulated genes showed enrichment of the hypothalamo-pituitary-thyroid ( HPT ) axis. Beta-estradiol was identified as an upstream regulator regardless of tissue. When LEPH and HEPH samples were compared, beta-estradiol was activated in HEPH in 3 of the 4 comparisons, which correlated to the number of beta-estradiol target genes up-regulated in HEPH. In in vitro pituitary cell cultures from LEPH and HEPH, thyroid hormone pretreatment negatively impacted gonadotropin subunit mRNA levels in cells from both LEPH and HEPH, with the effect being more prominent in HEPH cells. Additionally, the effect of estradiol pretreatment on gonadotropin subunit mRNA levels in HEPH cells was negative, whereas estradiol pretreatment increased gonadotropin subunit mRNA levels in LEPH cells. Conclusions: Up-regulation of the HPT axis in LEPH and upstream beta-estradiol activation in HEPH may play a role in regulating HPG axis function, and ultimately ovulation rates. Thyroid hormone and estradiol pretreatment impacted gonadotropin mRNA levels following GnRH stimulation, with the inhibitory effects of thyroid hormone more detrimental in HEPH and estradiol stimulatory effects more prominent in LEPH. Responsiveness to thyroid hormone and estradiol may be due to desensitization to thyroid hormone and estradiol in LEPH and HEPH, respectively, due to up-regulation of the HPT axis in LEPH and of the HPG axis in HEPH. Further studies will be necessary to identify possible target gene desensitization mechanisms and elicit the regulatory role of the HPT axis and beta-estradiol on ovulation rates in turkey hens.

CRISPR Knockdown of Kcnq3 Attenuates the M-current in NPY/AgRP Neurons

Arcuate nucleus Neuropeptide Y/Agouti-related peptide (NPY/AgRP) neurons drive ingestive behavior in response to the internal and external environment of an organism. NPY/AgRP neurons are adjacent to the median eminence, a circumventricular organ, and circulating metabolic factors and hormones communicate the energy state of the animal via these neurons by altering the excitability of NPY/AgRP neurons, which produces an appropriate change in behavior to maintain homeostasis. One example of this plasticity is seen in the M-current, a subthreshold, non-inactivating K+ current that acts to modulate excitability. Fasting decreases while estradiol increases the M-current through regulation of subunit mRNA expression of Kcnq 2, 3, & 5. KCNQ2/3 heteromers are thought to mediate the majority of the M-current. Here we used a recently developed single adeno-associated viral (AAV) vector containing a recombinase-dependent Staphylococcus aureus Cas9 (SaCas9) and a single guide RNA against Kcnq3 to selectively delete Kcnq3 in NPY/AgRP neurons to produce a loss of function in the M-current. We found that this virus was effective at knocking down Kcnq3 but not Kcnq2 expression. With the reduced KCNQ3 channel expression NPY/AgRP neurons were more depolarized, exhibited a higher input resistance, and the rheobase current needed to induce firing was significantly reduced, indicative of increased excitability. Although the resulting decrease in the M-current did not overtly alter ingestive behavior, it did significantly reduce the locomotor activity as measured in open field testing. Therefore, the SaCas9-sgKcnq3 is efficient to knock down Kcnq3 expression thereby reducing the M-current and increasing the excitability of NPY/AgRP neurons.

Transcriptome analysis of the hypothalamus and pituitary of turkey hens with low and high egg production

Background: High egg producing hens ( HEPH ) show increased hypothalamic and pituitary gene expression related to hypothalamo-pituitary-gonadal ( HPG ) axis stimulation as well as increased in vitro responsiveness to gonadotropin releasing hormone ( GnRH ) stimulation in the pituitary when compared to low egg producing hens ( LEPH ). Transcriptome analysis was performed on hypothalamus and pituitary samples from LEPH and HEPH to identify novel regulators of HPG axis function. Results: In the hypothalamus and pituitary, 4644 differentially expressed genes ( DEGs ) were identified between LEPH and HEPH, with 2021 genes up-regulated in LEPH and 2623 genes up-regulated in HEPH. In LEPH, up-regulated genes showed enrichment of the hypothalamo-pituitary-thyroid ( HPT ) axis. Beta-estradiol was identified as an upstream regulator regardless of tissue. When LEPH and HEPH samples were compared, beta-estradiol was activated in HEPH in 3 of the 4 comparisons, which correlated to the number of beta-estradiol target genes up-regulated in HEPH. In in vitro pituitary cell cultures from LEPH and HEPH, thyroid hormone pretreatment negatively impacted gonadotropin subunit mRNA levels in cells from both LEPH and HEPH, with the effect being more prominent in HEPH cells. Additionally, the effect of estradiol pretreatment on gonadotropin subunit mRNA levels in HEPH cells was negative, whereas estradiol pretreatment increased gonadotropin subunit mRNA levels in LEPH cells. Conclusions: Up-regulation of the HPT axis in LEPH and upstream beta-estradiol activation in HEPH may play a role in regulating HPG axis function, and ultimately ovulation rates. Thyroid hormone and estradiol pretreatment impacted gonadotropin mRNA levels following GnRH stimulation, with the inhibitory effects of thyroid hormone more detrimental in HEPH and estradiol stimulatory effects more prominent in LEPH. Responsiveness to thyroid hormone and estradiol may be due to desensitization to thyroid hormone and estradiol in LEPH and HEPH, respectively, due to up-regulation of the HPT axis in LEPH and of the HPG axis in HEPH. Further studies will be necessary to identify possible target gene desensitization mechanisms and elicit the regulatory role of the HPT axis and beta-estradiol on ovulation rates in turkey hens.

Transcriptome analysis of the hypothalamus and pituitary of turkey hens with low and high egg production

Background: High egg producing hens (HEPH) show increased hypothalamic and pituitary gene expression related to hypothalamo-pituitary-gonadal (HPG) axis stimulation as well as increased in vitro responsiveness to gonadotropin releasing hormone (GnRH) stimulation in the pituitary when compared to low egg producing hens (LEPH). Transcriptome analysis was performed on hypothalamus and pituitary samples from LEPH and HEPH to identify novel regulators of HPG axis function. Results: In the hypothalamus and pituitary, 4644 differentially expressed genes (DEGs) were identified between LEPH and HEPH, with 2021 genes up-regulated in LEPH and 2623 genes up-regulated in HEPH. In LEPH, up-regulated genes showed enrichment of the hypothalamo-pituitary-thyroid (HPT) axis. Beta-estradiol was identified as an upstream regulator regardless of tissue. When LEPH and HEPH samples were compared, beta-estradiol was activated in HEPH in 3 of the 4 comparisons, which correlated to the number of beta-estradiol target genes up-regulated in HEPH. In in vitro pituitary cell cultures from LEPH and HEPH, thyroid hormone pretreatment negatively impacted gonadotropin subunit mRNA levels in cells from both LEPH and HEPH, with the effect being more prominent in HEPH cells. Additionally, the effect of estradiol pretreatment on gonadotropin subunit mRNA levels in HEPH cells was negative, whereas estradiol pretreatment increased gonadotropin subunit mRNA levels in LEPH cells.Conclusions: Up-regulation of the HPT axis in LEPH and upstream beta-estradiol activation in HEPH may play a role in regulating HPG axis function, and ultimately ovulation rates. Furthermore, thyroid hormone and estradiol pretreatment impacted gonadotropin mRNA levels following GnRH stimulation, with the inhibitory effects of thyroid hormone being more detrimental in HEPH and estradiol stimulatory effects being more prominent in LEPH. Differential responsiveness to thyroid hormone and estradiol pretreatment may be due to desensitization of target genes to thyroid hormone and estradiol in LEPH and HEPH, respectively, in response general up-regulation of the HPT axis in LEPH and of the HPG axis in HEPH. Further studies will be necessary to identify possible target gene desensitization mechanisms and elicit the full role that the HPT axis and beta-estradiol upstream regulation play in egg production rates in turkey hens.

Integrin signaling: linking mechanical stimulation to skeletal muscle hypertrophy

The α7β1-integrin is a transmembrane adhesion protein that connects laminin in the extracellular matrix (ECM) with actin in skeletal muscle fibers. The α7β1-integrin is highly expressed in skeletal muscle and is concentrated at costameres and myotendious junctions, providing the opportunity to transmit longitudinal and lateral forces across the membrane. Studies have demonstrated that α7-integrin subunit mRNA and protein are upregulated following eccentric contractions as a mechanism to reinforce load-bearing structures and resist injury with repeated bouts of exercise. It has been hypothesized for many years that the integrin can also promote protein turnover in a manner that can promote beneficial adaptations with resistance exercise training, including hypertrophy. This review provides basic information about integrin structure and activation and then explores its potential to serve as a critical mechanosensor and activator of muscle protein synthesis and growth. Overall, the hypothesis is proposed that the α7β1-integrin can contribute to mechanical-load induced skeletal muscle growth via an mammalian target of rapamycin complex 1-independent mechanism.