strychnine and adenosine-3--5--cyclic-phosphorothioate

The cAMP-protein kinase A (PKA) pathway plays a critical role in regulating neuronal activity. Yet, how PKA signalling shapes the population activity of neurons that regulate respiratory rhythm and motor patterns in vivo is poorly defined. We determined the respiratory effects of focally inhibiting endogenous PKA activity in defined classes of respiratory neurons in the ventrolateral medulla and spinal cord by microinjection of the membrane-permeable PKA inhibitor Rp-adenosine 3',5'-cyclic monophosphothioate (Rp-cAMPS) in urethane-anaesthetized adult Sprague Dawley rats. Phrenic nerve activity, end-tidal CO2 and arterial pressure were recorded. Rp-cAMPS in the preBötzinger complex (preBötC) caused powerful, dose-dependent depression of phrenic burst amplitude and inspiratory period. Rp-cAMPS powerfully depressed burst amplitude in the phrenic premotor nucleus, but had no effect at the phrenic motor nucleus, suggesting a lack of persistent PKA activity here. Surprisingly, inhibition of PKA activity in the preBötC increased phrenic burst frequency, whereas in the Bötzinger complex phrenic frequency decreased. Pretreating the preBötC with strychnine, but not bicuculline, blocked the Rp-cAMPS-evoked increase in frequency, but not the depression of phrenic burst amplitude. We conclude that endogenous PKA activity in excitatory inspiratory preBötzinger neurons and phrenic premotor neurons, but not motor neurons, regulates network inspiratory drive currents that underpin the intensity of phrenic nerve discharge. We show that inhibition of PKA activity reduces tonic glycinergic transmission that normally restrains the frequency of rhythmic respiratory activity. Finally, we suggest that the maintenance of the respiratory rhythm in vivo is not dependent on endogenous cAMP-PKA signalling.

Topics: Animals; Bicuculline; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Glycine; Male; Phrenic Nerve; Protein Kinase Inhibitors; Rats; Rats, Sprague-Dawley; Respiratory Mechanics; Strychnine; Thionucleotides

Opioid receptors located on interneurons in the ventral tegmental area (VTA) inhibit GABA(A)-mediated synaptic transmission to dopamine projection neurons. The resulting disinhibition of dopamine cells in the VTA is thought to play a pivotal role in drug abuse; however, little is known about how this GABAA synapse is affected after chronic morphine treatment. The regulation of GABA release during acute withdrawal from morphine was studied in slices from animals treated for 6-7 d with morphine. Slices containing the VTA were prepared and maintained in morphine-free solutions, and GABAA IPSCs were recorded from dopamine cells. The amplitude of evoked IPSCs and the frequency of spontaneous miniature IPSCs measured in slices from morphine-treated guinea pigs were greater than placebo-treated controls. In addition, activation of adenylyl cyclase, with forskolin, and cAMP-dependent protein kinase, with Sp-cAMPS, caused a larger increase in IPSCs in slices from morphine-treated animals. Conversely, the kinase inhibitors staurosporine and Rp-CPT-cAMPS decreased GABA IPSCs to a greater extent after drug treatment. The results indicate that the probability of GABA release was increased during withdrawal from chronic morphine treatment and that this effect resulted from an upregulation of the cAMP-dependent cascade. Increased transmitter release from opioid-sensitive synapses during acute withdrawal may be one adaptive mechanism that results from prolonged morphine treatment.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Action Potentials; Adenylyl Cyclase Inhibitors; Adenylyl Cyclases; Animals; Colforsin; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Dopamine; Dopamine Antagonists; Enzyme Activation; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; GABA Antagonists; gamma-Aminobutyric Acid; Guinea Pigs; Interneurons; Morphine; Nerve Tissue Proteins; Organophosphorus Compounds; Patch-Clamp Techniques; Phorbol 12,13-Dibutyrate; Picrotoxin; Receptors, GABA-A; Salicylamides; Serotonin; Signal Transduction; Staurosporine; Strychnine; Substance Withdrawal Syndrome; Tegmentum Mesencephali; Thionucleotides