title: Distinct Calcium Sources Support Multiple Modes of Synaptic Release from Cranial Sensory Afferents.
abstract: Most craniosensory afferents have unmyelinated axons expressing TRP Vanilloid 1 (TRPV1) receptors in synaptic terminals at the solitary tract nucleus (NTS). Neurotransmission from these synapses is characterized by substantial asynchronous EPSCs following action potential-synched EPSCs and high spontaneous rates that are thermally sensitive. The present studies blocked voltage-activated calcium channels (CaV) using the nonselective CaV blocker Cd(2+) or the specific N-type blocker ω-conotoxin GVIA to examine the calcium dependence of the synchronous, asynchronous, spontaneous, and thermally gated modes of release. In rat brainstem slices containing caudal NTS, shocks to the solitary tract (ST) triggered synchronous ST-EPSCs and trailing asynchronous EPSCs. Cd(2+) or GVIA efficiently reduced both synchronous and asynchronous EPSCs without altering spontaneous or thermal-evoked transmission. Activation of TRPV1 with either the selective agonist resiniferatoxin (150 pm) or temperature augmented basal sEPSC rates but failed to alter the synchronous or asynchronous modes of release. These data indicate that calcium sourced through TRPV1 has no access to the synchronous or asynchronous release mechanism(s) and conversely that CaV-sourced calcium does not interact with the thermally evoked mode of release. Buffering intracellular calcium with EGTA-AM or BAPTA-AM reduced asynchronous EPSC rates earlier and to a greater extent than synchronous ST-EPSC amplitudes without altering sEPSCs or thermal sensitivity. Buffering therefore distinguishes asynchronous vesicles as possessing a highly sensitive calcium sensor located perhaps more distant from CaV than synchronous vesicles or thermally evoked vesicles from TRPV1. Together, our findings suggest separate mechanisms of release for spontaneous, asynchronous and synchronous vesicles that likely reside in unique, spatially separated vesicle domains.
  
  
  
  Most craniosensory fibers release glutamate using calcium entry from two sources: CaVs and TRPV1. We demonstrate that calcium segregation distinguishes three vesicle release mechanisms. Most surprisingly, asynchronous release is associated with CaV and not TRPV1 calcium entry. This reveals that asynchronous release is an additional and separate phenotypic marker of unmyelinated afferents rather than operated by TRPV1. The functional independence of the two calcium sources expands the regulatory repertoire of transmission and imbues these inputs with additional modulation targets for synaptic release not present at conventional CaV synapses. Peptides and lipid mediators may target one or both of these calcium sources at afferent terminals within the solitary tract nucleus to independently modify release from distinct, functionally segregated vesicle pools.
authors: Department of Physiology and Pharmacology, Oregon Health & Science University, Portland, Oregon 97239 fawley.jessica@gmail.com.
authors: Jessica A
authors: 0000-0002-8723-6096
authors: JA
authors: Fawley
authors: Department of Physiology and Pharmacology, Oregon Health & Science University, Portland, Oregon 97239.
authors: Mackenzie E
authors: 0000-0002-0469-3823
authors: ME
authors: Hofmann
authors: Department of Physiology and Pharmacology, Oregon Health & Science University, Portland, Oregon 97239.
authors: Michael C
authors: 0000-0003-0524-1172
authors: MC
authors: Andresen
mesh_terms: D000109:Acetylcholine / Q000378:metabolism; D000344:Afferent Pathways / Q000502:physiology*; D000704:Analysis of Variance; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D004224:Diterpenes / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004558:Electric Stimulation; D019706:Excitatory Postsynaptic Potentials / Q000187:drug effects / Q000502:physiology; D018698:Glutamic Acid / Q000378:metabolism; D066298:In Vitro Techniques; D008297:Male; D009474:Neurons / Q000502:physiology*; D018408:Patch-Clamp Techniques; D051381:Rats; D017207:Rats, Sprague-Dawley; D017552:Solitary Nucleus / Q000166:cytology; D013569:Synapses / Q000187:drug effects / Q000378:metabolism*; D050916:TRPV Cation Channels / Q000378:metabolism
chemical_list: D002121:Calcium Channel Blockers; D002614:Chelating Agents; D004224:Diterpenes; D050916:TRPV Cation Channels; C493922:Trpv1 protein, rat; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D018698:Glutamic Acid; D004533:Egtazic Acid; C024353:resiniferatoxin; D000109:Acetylcholine; D002118:Calcium
keywords: TRPV1; calcium; nucleus of the solitary tract; spontaneous release; synaptic transmission; unmyelinated