Photodynamic Modification of Native HCN Channels Expressed in Thalamocortical Neurons

Abstract

The photodynamic process requires three elements: light, oxygen, and photosensitizer, and involves the formation of singlet oxygen, the molecular oxygen in excited electronic states. Previously, we reported that heterologously expressed hyperpolarization-activated cAMP-gated (HCN) channels in excised membrane patches are sensitive to photodynamic modification (PDM). Here we extend this study to native HCN channels expressed in thalamocortical (TC) neurons in the ventrobasal (VB) complex of the thalamus and dopaminergic neurons (DA) of the ventral tegmental area (VTA). To do this, we introduced the photosensitizer FITC-cAMP into TCs or DAs of rodent brain slices via a whole-cell patch-clamp recording pipette. After illumination with blue light pulses, we observed an increase in the voltage-insensitive, instantaneous I inst component, accompanied by a long-lasting decrease in the hyperpolarization-dependent I h component. Both I h and the increased I inst after PDM could be blocked by the HCN blockers Cs + and ZD7288. When FITC and cAMP were dissociated and loaded into neurons as two separate chemicals, light application did not result in any long-lasting changes of the HCN currents. In contrast, light pulses applied to HCN2−/− neurons loaded with FITC-cAMP generated a much greater reduction in the I inst component compared to that of WT neurons. Next, we investigated the impact of the longlasting increases in I inst after PDM on the cellular physiology of VB neurons. Consistent with an upregulation of HCN channel function, PDM elicited a depolarization of the resting membrane potential (RMP). Importantly, Trolox-C, an effective quencher for singlet oxygen, could block the PDM-dependent increase in I inst and depolarization of the RMP. We propose that PDM of native HCN channels under physiological conditions may provide a photodynamic approach to alleviate HCN channelopathy in certain pathological conditions.

Effectiveness of Columbianadin, a Bioactive Coumarin Derivative, in Perturbing Transient and Persistent INa

“…As demonstrated in Figure 8 , the application of 10 µM CBN to the bath mildly lessened I h amplitude activated by 2 s step hyperpolarization from −40 to −120 mV, which was noticed to coincide with a lengthening in the activation time constant of I h . However, no change in the instantaneous amplitude of I h as recently reported [ 30 ] was detected in the presence of CBN. For example, when cells were exposed to 10 µM CBN, the I h amplitude measured at the end of 2-s hyperpolarizing pulse was mildly but significantly reduced by 9% ± 1% to 446 ± 29 pA ( n = 7, p < 0.05) from a control value of 492 ± 34 pA.…”

Photodynamic Activation of Cholecystokinin 1 Receptor Is Conserved in Mammalian and Avian Pancreatic Acini

“…Both CCK-stimulated and photodynamic activation of rat, mouse, and Peking duck CCK1R were confirmed by reversible inhibition by CCK1R antagonist devazepide ( Figure 1 , Figure 2 and Figure S1 ). The potential involvement of 1 O 2 in photodynamic CCK1R activation in rat, mouse, and Peking duck pancreatic acini was corroborated by complete inhibition by 1 O 2 quencher Trolox C ( Figure 1 , Figure 2 and Figure S1 ), which has been widely used to confirm the involvement of 1 O 2 in photodynamic modulation [ 45 , 46 , 47 ].…”

Dopaminergic amacrine cells express HCN channels in the developing and adult mouse retina

“…The former in the millimolar range can effectively block HCN channels from the extracellular side by blocking the ionconducting pore 34,35 . In contrast, ZD7288 must diffuse through the cell membrane to bind to the intracellular vestibule of the ion-conducting pore to block HCN channels 36 . Figure 5A illustrates an example of the blockade of the voltage sag of a DAC by 2 mM Cs + .…”