Microinjection of the alpha-subunit of the G protein Go2, but not Go1, reduces a voltage-sensitive calcium current.

Calcium currents can be modulated by receptor activation of the GTP-binding protein G(o). We have determined whether the two forms of G(o), Go1 and Go2, differentially regulate calcium current magnitude. Using identified neurons of the pond snail Helisoma, we demonstrate that a high-voltage-activated (HVA) calcium current is reduced by addition of the neuropeptide Phe-Met-Arg-Phe-amide (FMRFamide) and that this inhibition is mediated by a pertussis toxin (PTX)-sensitive G protein pathway. Using this calcium current as an assay for G protein activity, we microinjected GTP gamma S-activated alpha-subunits of G proteins into neuronal somata. We demonstrate that the calcium current is differentially regulated by the two forms of alpha o. Microinjection of alpha o2*, but not alpha o1*, reduces calcium current magnitude.

A relation between synaptic specificity and the acquisition of presynaptic properties.

1. The specificity of synaptogenesis of identified adult neurons of Helisoma was determined in cell culture. Cholinergic neuron B5 indiscriminately forms the presynaptic element of chemical connections with novel cholinoceptive target neurons and muscle. By contrast, cholinergic neuron B19 is selective and discriminates between novel and appropriate target cells. Neuron B19 forms chemical connections with appropriate muscle targets only. 2. The acquisition of presynaptic properties independent of target contact was studied for both identified neurons. Functional connections form between neuron B5 and novel targets within seconds of contact, indicating that this cell has synthesized the presynaptic apparatus before target contact. In contrast, neuron B19 showed no evidence of possessing the ability to release neurotransmitter. 3. To further study the development of presynaptic properties, a model system of giant synaptic terminals was developed. The soma of neuron B5, acutely isolated from the nervous system is non-secretory. In conditions that prevent the extension of neurites, somata gain the ability to release neurotransmitter. This experimentally tractable system was used to study the calcium currents of presynaptic neuron B5. Acutely-isolated non-secretory somata contain two types of calcium currents: low-voltage-activated (LVA) and high-voltage-activated (HVA). The types of calcium currents in the soma change when B5 gains its secretory capacity. Secretory somata contain HVA calcium current only. 4. Neuron B5 was also plated in conditions which permit the extension of neurites. LVA and HVA calcium currents were maintained in its soma (non-secretory) but HVA calcium current only was maintained in its growth cones (secretory). Thus, B5 differentially regulates the presence of specific calcium currents in its membrane in relation to local secretory capacity without target-derived cues. 5. These data suggest that neuron B5 has an intrinsic program which generates presynaptic calcium channels and secretory apparatus prior to target contact. This autonomy of initial presynaptic development may underlie the lack of target cell discrimination exhibited by B5 in synaptogenesis.