Key points Corticotroph cells of the anterior pituitary are electrically excitable and are an integral component of the hypothalamic‐pituitary‐adrenal axis which governs the neuroendocrine response to stress.Corticotrophs display predominantly single spike activity under basal conditions that transition to complex bursting behaviours upon stimulation by the hypothalamic secretagogues corticotrophin‐releasing hormone (CRH) and arginine vasopressin (AVP); however, the underlying mechanisms controlling bursting are unknown.In this study, we show that CRH and AVP induce different patterns of corticotroph electrical activity, and we use an electrophysiological approach combined with mathematical modelling to show the ionic mechanisms for these differential effects.The data reveal that secretagogue‐induced bursting is dependent on large conductance Ca2+‐activated K+ (BK) channels and is driven primarily by CRH whereas AVP promotes an increase in single‐spike frequency through BK‐independent pathways involving activation of non‐selective cation conductances.As corticotroph excitability is differentially regulated by CRH and AVP this may allow corticotrophs to respond appropriately to different stressors. AbstractAnterior pituitary corticotroph cells are a central component of the hypothalamic‐pituitary‐adrenal (HPA) axis essential for the neuroendocrine response to stress. Corticotrophs are excitable cells that receive input from two hypothalamic secretagogues, corticotrophin‐releasing hormone (CRH) and arginine vasopressin (AVP) to control the release of adrenocorticotrophic hormone (ACTH). Although corticotrophs are spontaneously active and increase in excitability in response to CRH and AVP the patterns of electrical excitability and underlying ionic conductances are poorly understood. In this study, we have used electrophysiological, pharmacological and genetic approaches coupled with mathematical modelling to investigate whether CRH and AVP promote distinct patterns of electrical excitability and to interrogate the role of large conductance calcium‐ and voltage‐activated potassium (BK) channels in spontaneous and secretagogue‐induced activity. We reveal that BK channels do not play a significant role in the generation of spontaneous activity but are critical for the transition to bursting in response to CRH. In contrast, AVP promotes an increase in single spike frequency, a mechanism independent of BK channels but dependent on background non‐selective conductances. Co‐stimulation with CRH and AVP results in complex patterns of excitability including increases in both single spike frequency and bursting. The ability of corticotroph excitability to be differentially regulated by hypothalamic secretagogues provides a mechanism for differential control of corticotroph excitability in response to different stressors.
Corticotroph cells from the anterior pituitary are an integral component of the hypothalamic-pituitary-adrenal (HPA) axis, which governs the neuroendocrine response to stress. Corticotrophs are electrically excitable and fire spontaneous single-spike action potentials and also display secretagogue-induced bursting behavior. The HPA axis function is dependent on effective negative feedback in which elevated plasma glucocorticoids result in inhibition at the level of both the pituitary and the hypothalamus. In this study, we have used an electrophysiological approach coupled with mathematical modeling to investigate the regulation of spontaneous and CRH/arginine vasopressin-induced activity of corticotrophs by glucocorticoids. We reveal that pretreatment of corticotrophs with 100 nM corticosterone (CORT; 90 and 150 min) reduces spontaneous activity and prevents a transition from spiking to bursting after CRH/arginine vasopressin stimulation. In addition, previous studies have identified a role for large-conductance calcium- and voltage-activated potassium (BK) channels in the generation of secretagogue-induced bursting in corticotrophs. Using the dynamic clamp technique, we demonstrated that CRH-induced bursting can be switched to spiking by subtracting a fast BK current, whereas the addition of a fast BK current can induce bursting in CORT-treated cells. In addition, recordings from BK knockout mice (BK−/−) revealed that CORT can also inhibit excitability through BK-independent mechanisms to control spike frequency. Thus, we have established that glucocorticoids can modulate multiple properties of corticotroph electrical excitability through both BK-dependent and BK-independent mechanisms.
Key points r Corticotroph cells of the anterior pituitary are electrically excitable and are an integral component of the hypothalamic-pituitary-adrenal axis which governs the neuroendocrine response to stress. r Corticotrophs display predominantly single spike activity under basal conditions that transition to complex bursting behaviours upon stimulation by the hypothalamic secretagogues corticotrophin-releasing hormone (CRH) and arginine vasopressin (AVP); however, the underlying mechanisms controlling bursting are unknown. r In this study, we show that CRH and AVP induce different patterns of corticotroph electrical activity, and we use an electrophysiological approach combined with mathematical modelling to show the ionic mechanisms for these differential effects. r The data reveal that secretagogue-induced bursting is dependent on large conductance Ca 2+-activated K + (BK) channels and is driven primarily by CRH whereas AVP promotes an increase in single-spike frequency through BK-independent pathways involving activation of non-selective cation conductances. r As corticotroph excitability is differentially regulated by CRH and AVP this may allow cortico-trophs to respond appropriately to different stressors. Abstract Anterior pituitary corticotroph cells are a central component of the hypothalamic-pituitary-adrenal (HPA) axis essential for the neuroendocrine response to stress. Corticotrophs are excitable cells that receive input from two hypothalamic secretagogues, corticotrophin-releasing hormone (CRH) and arginine vasopressin (AVP) to control the release of adrenocorticotrophic hormone (ACTH). Although corticotrophs are spontaneously active and increase in excitability in response to CRH and AVP the patterns of electrical excitability and underlying ionic conductances are poorly understood. In this study, we have used electrophysiological, pharmacological and genetic approaches coupled with mathematical modelling to investigate whether CRH and AVP promote distinct patterns of electrical excitability and to interrogate the role of large conductance calcium-and voltage-activated potassium (BK) channels in spontaneous and secretagogue-induced activity. We reveal that BK channels do not play a significant role in the generation of spontaneous activity but are critical for the transition to bursting in response to CRH. In contrast, AVP promotes an increase in single spike frequency, a mechanism independent of BK channels but dependent on background non-selective conductances. Co-stimulation with CRH and AVP results in complex patterns of excitability including increases in both single spike frequency and bursting. The ability of corticotroph excitability to be differentially regulated by hypothalamic secretagogues provides a mechanism for differential control of corticotroph excitability in response to different stressors.
The properties and physiological function of pore-forming α-subunits of large conductance calcium- and voltage-activated potassium (BK) channels are potently modified by their functional coupling with regulatory subunits in many tissues. However, mechanisms that might control functional coupling are very poorly understood. Here we show that S -acylation, a dynamic post-translational lipid modification of proteins, of the intracellular S0–S1 loop of the BK channel pore-forming α-subunit controls functional coupling to regulatory β1-subunits. In HEK293 cells, α-subunits that cannot be S -acylated show attenuated cell surface expression, but expression was restored by co-expression with the β1-subunit. However, we also found that nonacylation of the S0–S1 loop reduces functional coupling between α- and β1-subunits by attenuating the β1-subunit-induced left shift in the voltage for half-maximal activation. In mouse vascular smooth muscle cells expressing both α- and β1-subunits, BK channel α-subunits were endogenously S -acylated. We further noted that S -acylation is significantly reduced in mice with a genetic deletion of the palmitoyl acyltransferase (Zdhhc23) that controls S -acylation of the S0–S1 loop. Genetic deletion of Zdhhc23 or broad-spectrum pharmacological inhibition of S -acylation attenuated endogenous BK channel currents independently of changes in cell surface expression of the α-subunit. We conclude that functional effects of S -acylation on BK channels depend on the presence of β1-subunits. In the absence of β1-subunits, S -acylation promotes cell surface expression, whereas in its presence, S -acylation controls functional coupling. S -Acylation thus provides a mechanism that dynamically regulates the functional coupling with β1-subunits, enabling an additional level of conditional, cell-specific control of ion-channel physiology.
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