Role of mitochondrial dysfunction in the Ca\u3csup\u3e2+\u3c/sup\u3e-induced decline of transmitter release at K\u3csup\u3e+\u3c/sup\u3e-depolarized motor neuron terminals

The present study tested whether a Ca -induced disruption of mitochondrial function was responsible for the decline in miniature endplate current (MEPC) frequency that occurs with nerve-muscle preparations maintained in a 35 mM potassium propionate (35 mM KP) solution containing elevated calcium. When the 35 mM KP contained control Ca (1 mM), the MEPC frequency increased and remained elevated for many hours, and the mitochondria within twitch motor neuron terminals were similar in appearance to those in unstimulated terminals. All nerve terminals accumulated FM1-43 when the dye was present for the final 6 min of a 300-min exposure to 35 mM KP with control Ca . In contrast, when Ca was increased to 3.6 mM in the 35 mM KP solution, the MEPC frequency initially reached frequencies \u3e350 s but then gradually fell approaching frequencies \u3c50 s . A progressive swelling and eventual distortion of mitochondria within the twitch motor neuron terminals occurred during prolonged exposure to 35 mM KP with elevated Ca . After ~300 min in 35 mM KP with elevated Ca , only 58% of the twitch terminals accumulated FM1-43. The decline in MEPC frequency in 35 mM KP with elevated Ca was less when 15 mM glucose was present or when preparations were pretreated with 10 μM oligomycin and then bathed in the 35 mM KP with glucose. When glucose was present, with or without oligomycin pretreatment, a greater percentage of twitch terminals accumulated FM1-43. However, the mitochondria in these preparations were still greatly swollen and distorted. We propose that prolonged depolarization of twitch motor neuron terminals by 35 mM KP with elevated Ca produced a Ca - induced decrease in mitochondrial ATP production. Under these conditions, the cytosolic ATP/ADP ratio was decreased thereby compromising both transmitter release and refilling of recycled synaptic vesicles. The addition of glucose stimulated glycolysis which contributed to the maintenance of required ATP levels. 2+ 2+ 2+ 2+ -1 -1 2+ 2+ 2+ 2+ 2

Organization of Corticostriatal Projections From the Vibrissal Representations in the Primary Motor and Somatosensory Cortical Areas of Rodents

To characterize corticostriatal projections from rodent sensorimotor cortex, the anterograde tracers biotinylated dextran amine (BDA) and fluororuby (FR) were injected into the whisker representations of the primary motor (MI) and somatosensory (SI) cortices. Reconstructions of labeled terminals and their beaded varicosities in the neostriatum and thalamus were analyzed quantitatively to determine the degree of labeled overlap in both of these subcortical structures. Corticostriatal projections from the vibrissal representation in MI were more extensive than corresponding projections from SI. Both cortical areas sent dense projections to the dorsolateral neostriatum, but the MI vibrissal representation also projected to regions located more rostrally and medially. Despite these differences, both MI and SI projected to overlapping parts of the dorsolateral neostriatum. Tracer injections in both cortical areas also produced dense anterograde and retrograde labeling in the medial sector of the posterior complex of the thalamus (POm). Because POm is somatotopically organized and has reciprocal connections with both SI and MI cortices, the amount of labeled overlap in POm was used to indicate whether the tracers were injected into corresponding whisker representations of MI and SI. We found that the proportion of labeled overlap in the neostriatum was highly correlated with the amount of labeled overlap in POm. These results indicate that the rodent neostriatum receives convergent projections from corresponding regions in MI and SI cortex. Furthermore, the thalamocortical projections of the POm indicate that it may modulate corticostriatal outputs from corresponding representations in MI and SI

PACAP peptides modulate guinea pig cardiac neuron membrane excitability and neuropeptide expression

Morphological studies identified PACAP-immunoreactive nerve fibers in dense pericellular arrangements around virtually every cholinergic parasympathetic neuron of guinea pig cardiac ganglia; all postganglionic cardiac neurons expressed membrane-associated PAC receptor protein. Characterization of the alternative splice variants established predominant expression of the PAC (very short) receptor transcript containing neither HIP nor HOP exons. PACAP depolarized cardiac neurons and increased membrane excitability; the excitability resulted from neither altered action potential properties nor inhibition of I . Treatment of cardiac ganglia explants with PACAP significantly reduced the numbers of cholinergic neurons coexpressing somatostatin immunoreactivity, which did not appear to be correlated with prosomatostatin mRNA expression. The PACAP-mediated decrease in somatostatin immunoreactive neurons required calcium influx through L-type calcium channels and activation of adenylyl cyclase, whereas activation of phospholipase C or protein kinase A was not required. These observations indicate that PACAP through the PAC receptors elicits complex actions on guinea pig parasympathetic cardiac ganglia neurons, including modulation of membrane ion conductances and modulation of neuropeptide expression. 1 1 M

Presynaptic function is altered in snake K+-depolarized motor nerve terminals containing compromised mitochondria

Presynaptic function was investigated at K+-stimulated motor nerve terminals in snake costocutaneous nerve muscle preparations exposed to carbonyl cyanide m-chlorophenylhydrazone (CCCP, 2 μm), oligomycin (8 μg ml−1) or CCCP and oligomycin together.Miniature endplate currents (MEPCs) were recorded at -150 mV with two-electrode voltage clamp. With all three drug treatments, during stimulation by elevated K+ (35 mm), MEPC frequencies initially increased to values > 350 s−1, but then declined. The decline occurred more rapidly in preparations treated with CCCP or CCCP and oligomycin together than in those treated with oligomycin alone.Staining with FM1-43 indicated that synaptic vesicle membrane endocytosis occurred at some CCCP- or oligomycin-treated nerve terminals after 120 or 180 min of K+ stimulation, respectively.The addition of glucose to stimulate production of ATP by glycolysis during sustained K+ stimulation attenuated the decline in MEPC frequency and increased the percentage of terminals stained by FM1-43 in preparations exposed to either CCCP or oligomycin.We propose that the decline in K+-stimulated quantal release in preparations treated with CCCP, oligomycin or CCCP and oligomycin together could result from a progressive elevation of intracellular calcium concentration ([Ca2+]i). For oligomycin-treated nerve terminals, a progressive elevation of [Ca2+]i could occur as the cytoplasmic ATP/ADP ratio decreases, causing energy-dependent Ca2+ buffering mechanisms to fail. The decline in MEPC frequency could occur more rapidly in preparations treated with CCCP or CCCP and oligomycin together because mitochondrial Ca2+ buffering and ATP production were both inhibited. Therefore, the proposed sustained elevation of [Ca2+]i could occur more rapidly