In vitro characterization of T-type calcium channel antagonist TTA-A2 and in vivo effects on arousal in mice.
T-type calcium channels have been implicated in many behaviorally important neurophysiological processes, and altered channel activity has been linked to the pathophysiology of neurological disorders such as insomnia, epilepsy, Parkinson's disease, depression, schizophrenia, and pain. We have previously identified a number of potent and selective T-type channel antagonists (Barrow et al., 2007; Shipe et al., 2008; Yang et al., 2008). Here we describe the properties of the antagonist TTA-A2 [2-(4-cyclopropylphenyl)-N-((1R)-1-{5-[(2,2,2-trifluoroethyl)oxo]-pyridin-2-yl}ethyl)acetamide], assessed in patch-clamp experiments. TTA-A2 blocks T-type channels (Ca(v)3.1, 3.2, 3.3) voltage dependently and with high potency (IC(50) ∼100 nM). Stimulation at 3 Hz revealed additional use dependence of inhibition. A hyperpolarized shift of the channel availability curve and delayed channel recovery from inactivation suggest that the compound preferentially interacts with and stabilizes inactivated channels. The compound showed a ∼300-fold selectivity for Ca(v)3 channels over high-voltage activated calcium channels. Inhibitory effects on native T-type currents were confirmed in brain slice recordings from the dorsal lateral geniculate nucleus and the subthalamic nucleus. Furthermore, we demonstrate that in vivo T-type channel inhibition by TTA-A2 suppresses active wake and promotes slow-wave sleep in wild-type mice but not in mice lacking both Ca(v)3.1 and Ca(v)3.3, suggesting the selective effect of TTA-A2 on recurrent thalamocortical network activity. The discovery of the potent and selective T-type channel antagonist TTA-A2 has enabled us to study the in vivo effects of pharmacological T-channel inhibition on arousal in mice, and it will help to explore the validity of these channels as potential drug targets for sleep-related and other neurological diseases.
Kraus
Richard L
RL
Department of Neuroscience, Merck Research Laboratories, 770 Sumneytown Pike, West Point, PA 19486, USA. richard_kraus@merck.com
Li
Yuxing
Y
Gregan
Yun
Y
Gotter
Anthony L
AL
Uebele
Victor N
VN
Fox
Steven V
SV
Doran
Scott M
SM
Barrow
James C
JC
Yang
Zhi-Qiang
ZQ
Reger
Thomas S
TS
Koblan
Kenneth S
KS
Renger
John J
JJ
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D001143:Arousal / Q000187:drug effects*; D044482:Benzeneacetamides / Q000737:chemistry / Q000494:pharmacology* / Q000627:therapeutic use; D002121:Calcium Channel Blockers / Q000737:chemistry / Q000494:pharmacology* / Q000627:therapeutic use; D020747:Calcium Channels, T-Type / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D005829:Geniculate Bodies / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D015394:Molecular Structure; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D018408:Patch-Clamp Techniques; D011725:Pyridines / Q000737:chemistry / Q000494:pharmacology* / Q000627:therapeutic use; D051381:Rats; D017207:Rats, Sprague-Dawley; D020921:Sleep Arousal Disorders / Q000188:drug therapy / Q000378:metabolism
C553486:2-(4-cyclopropylphenyl)-N-(1-(5-((2,2,2-trifluoroethyl)oxo)pyridin-2-yl)ethyl)acetamide; D044482:Benzeneacetamides; D002121:Calcium Channel Blockers; D020747:Calcium Channels, T-Type; D011725:Pyridines